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Sample records for interleukin-10 gene transfer

  1. Association of interleukin 10 and transforming growth factor β gene polymorphisms with chronic idiopathic urticaria.

    PubMed

    Tavakol, Marzieh; Movahedi, Masoud; Amirzargar, Ali Akbar; Aryan, Zahra; Bidoki, Alireza Zare; Heidari, Kimia; Soltani, Samaneh; Gharagozlou, Mohammad; Aghamohammadi, Asghar; Nabavi, Mohammad; Nasiri, Rasoul; Ahmadvand, Alireza; Rezaei, Nima

    2014-01-01

    Transforming growth factor β (TGF-β) and interleukin 10 (IL-10) are two anti-inflammatory cytokines that are implicated in the pathogenesis of urticaria. The goal of this study was to examine the possible association of polymorphisms of TGF-β and IL-10 genes with susceptibility to chronic idiopathic urticaria (CIU). This study was conducted on 90 patients with CIU. Polymerase chain reaction (PCR) was done to determine the genotype at 5 polymorphic sites; TGF-β (codon10C/T and codon25G/C) and IL-10 (-1082G/A, -819C/T, and -592C/A). The C allele at codon 25 of TGF-β was more prevalent in CIU patients compared to controls (OR = 9.5, 95% CI = 5.4-16.8, P<0.001). Genotypes of CT and CG at 10 and 25 codons of TGF-β gene, respectively, and AG, CT, and CA for loci of -1082, -819, and -592 of IL-10 gene were significantly higher in CIU patients (P<0.001). In haplotype analysis, frequency of TGF-β haplotypes differed between patients with CIU and controls; CC haplotype was overrepresented, while CG and TG haplotypes were underrepresented (P<0.001). These results suggest that TGF-β and IL-10 genetic variability could contribute to susceptibility to CIU. Additionally, patients with CIU seem to have genotypes leading to high production of TGF-β and IL-10. PMID:25580782

  2. Interleukin-10 -1082 G/A gene polymorphisms in Egyptian children with CAP

    PubMed Central

    Azab, Seham F.; Abdalhady, Mohamed A.; Elsaadany, Hosam F.; Elkomi, Mohamed A.; Elhindawy, Eman M.; Sarhan, Dina T.; Salam, Mohamed M.A.; Allah, Mayy A.N.; Emam, Ahmed A.; Noah, Maha A.; Abdelsalam, Nasser I.; Abdellatif, Sawsan H.; Rass, Anwar A.; Ismail, Sanaa M.; Gheith, Tarek; Aziz, Khalid A.; Hamed, Mohammed E.; Abdelrahman, Hind M.; Ahmed, Ahmed R.; Nabil, Rehab M.; Abdulmaksoud, Rehab S.; Yousef, Hala Y.

    2016-01-01

    Abstract Community-acquired pneumonia (CAP) is one of the leading causes of death worldwide. Cytokines are involved in the pathogenesis of CAP. To date, only a few studies concerned the association of interleukin-10 (IL-10) gene polymorphisms with CAP. In this study, we aimed to investigate whether the -1082(G/A) polymorphism in the promoter region of the IL-10 gene is involved in susceptibility to and the outcome of CAP, and we also measured the serum level of IL-10 to assess its relation to such polymorphism. This was a case–control study included 100 patients with CAP, and matched with age, gender, and ethnicity of 100 healthy control children. IL-10 -1082(G/A) gene polymorphism was genotyped by polymerase chain reaction-restriction fragment length polymorphism, while the serum IL-10 levels were measured by ELISA method. Compared to the controls subjects, the frequencies of the IL-10 -1082 AA genotype and A allele were observed to be overrepresented in patients with CAP (51%; odds ratio [OR] = 2.8; 95% confidence interval [CI]: 1.5–5.3 for the AA genotype; P < 0.01) and (70%; OR: 1.95; 95% CI: 1.27–3.00 for the A allele; P < 0.01, respectively). We found that patients with the GG genotype had significantly higher serum IL-10 levels (46.7 ± 9.5 pg/mL) compared to those with AG genotype (21.8 ± 4.5 pg/mL) and AA genotype (11.5 ± 3.3 pg/mL); P < 0.01, respectively. Our data revealed a significant positive association between the -1082 GG genotype and susceptibility to severe sepsis, acute respiratory failure, and hospital mortality (OR: 3.8; 95% CI: 1.3–11.2; P < 0.01). We demonstrate for the first time, to the best of our knowledge, that IL-10 -1082 (G/A) gene polymorphism may contribute to susceptibility to CAP in Egyptian children. Moreover, we observed that the presence of a G allele or GG genotype at the -1082 position of the promoter region of the IL-10 gene constitute risk factors for developing severe sepsis

  3. Interleukin-10 Gene Polymorphisms are Associated With Freedom From Treatment Failure for Patients With Hodgkin Lymphoma

    PubMed Central

    Schoof, Nils; Franklin, Jeremy; Fürst, Robert; Zander, Thomas; von Bonin, Frederike; Peyrade, Frederic; Trümper, Lorenz; Diehl, Volker; Engert, Andreas

    2013-01-01

    Background. Hodgkin lymphoma (HL) is a lymphoid malignancy characterized by the production of various cytokines possibly involved in immune deregulation. Interleukin-10 (IL-10) serum levels have been associated with clinical outcome in patients with HL. Because host genetic variations are known to alter the expression and function of cytokines and their receptors, we investigated whether genetic variations influence clinical outcome of patients with HL. Methods. A total of 301 patients with HL who were treated within randomized trials by the German Hodgkin Study Group were included in this exploratory retrospective study. Gene variations of IL-10 (IL-10-597AC, rs1800872; IL-10-824CT, rs1800871; IL-10-1087AG, rs1800896; IL-10-3538AT, rs1800890; IL-10-6208CG, rs10494879; IL-10-6752AT, rs6676671; IL-10-7400InDel), IL-13 (IL-13-1069CT, rs1800925; IL-13Q144R, rs20541), and IL-4R (IL-4RI75V, rs1805010; IL-4RQ576R, rs1801275) were genotyped. Results. Inferior freedom from treatment failure (FFTF) was found in patients harboring the IL-10-597AA, IL-10-824TT, or the IL-10-1087AA genotype. In contrast, the IL-10-1087G-824C-597C haplotype present in about 48% of analyzed HL patients is nominally significant for a better FFTF in a Cox-Regression model accounting for stage and treatment. No associations were observed between the other IL-10 gene variations, IL-13-1069CT, IL-13Q144R, IL-4RI75V, IL-4RQ576R and the clinical outcome of patients with HL. Conclusions. Our study provides further evidence that proximal IL-10 promoter gene variations are associated with clinical course of patients with HL. However, treatment success and survival rates are already at a very high rate, supporting the need to design studies focusing on identification of predictors to reduce the side effects of therapy. PMID:23299779

  4. ACC interleukin-10 gene promoter haplotype as a breast cancer risk factor predictor among Jordanian females

    PubMed Central

    Atoum, Manar Fayiz

    2016-01-01

    Introduction Interleukin-10 (IL-10) is a multifactorial cytokine with a complex biological role in breast cancer. The aims of this study were to investigate any association between IL-10 gene promoter polymorphisms, 1082A>/G, −819T>C, and −592A>C, or haplotypes and breast cancer risk among Jordanian women and to evaluate any association between the most common haplotype with clinicopathological features of breast cancer. Patients and methods A total of 202 breast cancer patients and 210 age-matched healthy control subjects were genotyped for −1082A/G, −819T/C, and −592A/C single nucleotide polymorphisms in the promoter region of the IL-10 gene by polymerase chain reaction-restriction fragment length polymorphism. Study patients and control subjects were recruited from Prince Hamzah Hospital, Amman, Jordan (2012–2013). Ethical approval and signed consent forms were signed by all participants. DNA was extracted, and polymerase chain reaction fragments were amplified and restriction digested by MnII, MaeIII, and RsaI. Results This study showed no statistically significant difference between −1082A/G, −819T/C, and −592A/C IL-10 genotypes or alleles among breast cancer patients or controls. Four different haplotypes ATA, ACC, GTA, and ACA within the IL-10 promoter gene were determined among both breast cancer and control groups. The most frequent haplotype was ACC among breast cancer patients and controls (41.6% and 40.7%, respectively). No statistical differences in these haplotypes among breast cancer patients or controls were determined. Analysis of the most common ACC haplotype showed statistical difference in positive estrogen receptor (P=0.022), positive progesterone receptor (P=0.004), cancer grade (P=0.0001), and cancer stage (P=0.009) among the ACC haplotype compared to non-ACC haplotype. Conclusion To our knowledge, this is the first report studying the association of IL-10 haplotype with breast cancer risk events among Jordanian females. The

  5. Co-transfection of decorin and interleukin-10 modulates pro-fibrotic extracellular matrix gene expression in human tenocyte culture

    PubMed Central

    Abbah, Sunny A.; Thomas, Dilip; Browne, Shane; O’Brien, Timothy; Pandit, Abhay; Zeugolis, Dimitrios I.

    2016-01-01

    Extracellular matrix synthesis and remodelling are driven by increased activity of transforming growth factor beta 1 (TGF-β1). In tendon tissue repair, increased activity of TGF-β1 leads to progressive fibrosis. Decorin (DCN) and interleukin 10 (IL-10) antagonise pathological collagen synthesis by exerting a neutralising effect via downregulation of TGF-β1. Herein, we report that the delivery of DCN and IL-10 transgenes from a collagen hydrogel system supresses the constitutive expression of TGF-β1 and a range of pro-fibrotic extracellular matrix genes. PMID:26860065

  6. Co-transfection of decorin and interleukin-10 modulates pro-fibrotic extracellular matrix gene expression in human tenocyte culture

    NASA Astrophysics Data System (ADS)

    Abbah, Sunny A.; Thomas, Dilip; Browne, Shane; O'Brien, Timothy; Pandit, Abhay; Zeugolis, Dimitrios I.

    2016-02-01

    Extracellular matrix synthesis and remodelling are driven by increased activity of transforming growth factor beta 1 (TGF-β1). In tendon tissue repair, increased activity of TGF-β1 leads to progressive fibrosis. Decorin (DCN) and interleukin 10 (IL-10) antagonise pathological collagen synthesis by exerting a neutralising effect via downregulation of TGF-β1. Herein, we report that the delivery of DCN and IL-10 transgenes from a collagen hydrogel system supresses the constitutive expression of TGF-β1 and a range of pro-fibrotic extracellular matrix genes.

  7. Interleukin-10 Promoter Gene Polymorphisms and Susceptibility to Tuberculosis: A Meta-Analysis

    PubMed Central

    Gao, Xuan; Chen, Junjun; Tong, Zhongkai; Yang, Guangdie; Yao, Yinan; Xu, Fei; Zhou, Jianying

    2015-01-01

    Objective As an update to other recent meta-analyses, the purpose of this study was to explore whether interleukin-10 (IL-10) polymorphisms and their haplotypes contribute to tuberculosis (TB) susceptibility. Methods We searched for published case-control studies examining IL-10 polymorphisms and TB in PubMed, EMBASE, Cochrane Central Register of Controlled Trials (CENTRAL), Wanfang databases and the Chinese National Knowledge Infrastructure (CNKI). Odds ratios (ORs) with 95% confidence intervals (CIs) were used to calculate the strengths of the associations. Results A total of 28 studies comprising 8,242 TB patients and 9,666 controls were included in the present study. There were no significant associations between the -1082G/A, -819C/T, and -592A/C polymorphisms and TB in the pooled samples. Subgroup analyses revealed that the -819T allele was associated with an increased TB risk in Asians in all genetic models (T vs. C: OR=1.17, 95% CI=1.05-1.29, P=0.003; TT vs. CC: OR=1.37, 95% CI=1.09-1.72, P=0.006; CT+TT vs. CC: OR=1.33, 95% CI=1.09-1.63, P=0.006; TT vs. CT+CC: OR=1.17, 95% CI=1.02-1.35, P=0.03) and that the -592A/C polymorphism was significantly associated with TB in Europeans under two genetic models (A vs. C: OR=0.77, 95% CI=0.60-0.98, P=0.03; AA vs. CC: OR=0.53, 95% CI=0.30-0.95, P=0.03). Furthermore, the GCC IL-10 promoter haplotype was associated with an increased risk of TB (GCC vs. others: P=1.42, 95% CI=1.02-1.97, P=0.04). Subgroup analyses based on ethnicity revealed that the GCC haplotype was associated with a higher risk of TB in Europeans, whereas the ACC haplotype was associated with a lower TB risk in both Asians and Europeans. Conclusions This meta-analysis suggests that the IL-10-819T/C polymorphism is associated with the risk of TB in Asians and that the IL-10-592A/C polymorphism may be a risk factor for TB in Europeans. Furthermore, these data indicate that IL-10 promoter haplotypes play a vital role in the susceptibility to or protection

  8. Intrathecal polymer-based interleukin-10* gene delivery for neuropathic pain

    PubMed Central

    MILLIGAN, ERIN D.; SODERQUIST, RYAN G.; MALONE, STEPHANIE M.; MAHONEY, JOHN H.; HUGHES, TRAVIS S.; LANGER, STEPHEN J.; SLOANE, EVAN M.; MAIER, STEVEN F.; LEINWAND, LESLIE A.; WATKINS, LINDA R.; MAHONEY, MELISSA J.

    2007-01-01

    Research on communication between glia and neurons has increased in the past decade. The onset of neuropathic pain, a major clinical problem that is not resolved by available therapeutics, involves activation of spinal cord glia through the release of proinflammatory cytokines in acute animal models of neuropathic pain. Here, we demonstrate for the first time that the spinal action of the proinflammatory cytokine, interleukin 1 (IL-1) is involved in maintaining persistent (2 months) allodynia induced by chronic-constriction injury (CCI). The anti-inflammatory cytokine IL-10 can suppress proinflammatory cytokines and spinal cord glial amplification of pain. Given that IL-1 is a key mediator of neuropathic pain, developing a clinically viable means of long-term delivery of IL-10 to the spinal cord is desirable. High doses of intrathecal IL-10-gene therapy using naked plasmid DNA (free pDNA-IL-10) is effective, but the dose required limits its potential clinical utility. Here we show that intrathecal gene therapy for neuropathic pain is improved sufficiently using two, distinct synthetic polymers, poly(lactic-co-glycolic) and polyethylenimine, that substantially lower doses of pDNA-IL-10 are effective. In conclusion, synthetic polymers used as i.t. gene-delivery systems are well-tolerated and improve the long-duration efficacy of pDNA-IL-10 gene therapy. PMID:18079973

  9. Interferon-γ and Interleukin-10 Gene Polymorphisms are not Predictors of Chronic Hepatitis C (Genotype-4) Disease Progression.

    PubMed

    Bahgat, Nermine Ahmed; Kamal, Manal Mohamed; Abdelaziz, Ashraf Omar; Mohye, Mohamed Ahmed; Shousha, Hend Ibrahim; ahmed, Mae Mohamed; Elbaz, Tamer Mahmoud; Nabil, Mohamed Mahmoud

    2015-01-01

    Immunoregulatory cytokines have an influence on hepatitis C virus (HCV) infection outcome. This study aimed to determine whether single nucleotide polymorphisms (SNP) in IFN- γ and IL-10 genes are associated with susceptibility and/or are markers of prognosis regarding chronic hepatitis C outcomes. IFN γ (+874T/A) and IL-10 (-1082G/A) genotypes were determined in 75 HCV genotype 4 patients with different disease severities (chronic hepatitis, n=25, liver cirrhosis and hepatocellular carcinoma (HCC) on top of liver cirrhosis, n=50) and 25 healthy participants using allele-specific polymerase chain reaction. No statistical differences in allele or genotype distributions of IFN γ and IL-10 genes were detected between patients and controls or between patientgroups. No significant difference in the frequency of IL-10 SNP at position -1082 or IFN-γ at position +874T/A was found between chronic HCV genotype 4 and with progression of disease severity in liver cirrhosis or HCC. In conclusion; interferon-γ and interleukin-10 gene polymorphisms are not predictors of disease progression in patients with chronic hepatitis C (Genotype-4). PMID:26163635

  10. Evaluating the association of interleukin-10 gene promoter -592 A/C polymorphism with lupus nephritis susceptibility

    PubMed Central

    Abdallah, Emad; Waked, Emam; Abdelwahab, Mahmoud A.

    2015-01-01

    Background Interleukin-10 (IL-10) is an important immunoregulatory cytokine. There are few studies evaluating the association between IL-10 and lupus nephritis (LN). The aim of this study was to evaluate the association of IL-10 gene promoter -592 A/C with LN susceptibility. Methods The study was conducted on 84 patients with systemic lupus erythematosus (SLE). Patients were divided into LN group (Group I, 48 patients) and non-LN group (Group II, 36 patients). The -592 A/C polymorphisms in IL-10 promoter gene were determined by polymerase chain reaction and restriction fragment length polymorphism in both groups. IL-10 was determined by enzyme-linked immunosorbent assay. Frequencies of the genotypes were compared between LN and non-LN patients and among LN patients with different pathologic classes. Results There was a significant increase in serum level of IL-10 (P = 0.001) in Group I compared with Group II and significant positive correlation between serum IL-10 and SLE disease activity index (r = 0.466, P = 0.001) in Group I. There were no significant differences in the distribution of the IL-10 gene promoter -592 A/C genotypes or the allele frequencies between Groups I and II. There was no significant difference between AC/CC and AA genotypes with SLE disease activity index, proteinuria, hematuria, anti-double-stranded DNA, and IL-10 in Group I. There was no significant difference in the distribution of AC and CC genotypes among different pathologic LN classes. Conclusion IL-10 suggested to play a role in pathogenesis and development of LN. However, the promoter -592 A/C of IL-10 gene suggested to be not associated with serum IL-10 levels or LN susceptibility. In addition, it appears that promoter -592 A/C of IL-10 gene not associated with LN activity or the pathologic classes of LN. PMID:27069855

  11. Interleukin 10 gene promoter polymorphisms in women with early-onset pre-eclampsia.

    PubMed

    Sowmya, S; Sri Manjari, K; Ramaiah, A; Sunitha, T; Nallari, P; Jyothy, A; Venkateshwari, A

    2014-11-01

    Pre-eclampsia is one of the most serious disorders of human pregnancy and T helper type 1 (Th1)/Th2 imbalance plays a major role in its aetiology. The Th2 cytokine, interleukin (IL)-10, plays a significant role in the maintenance of pregnancy. The present study is aimed at understanding the role of IL-10 promoter polymorphisms (-1082 G/A; -592 A/C and -819 C/T) and their haplotypes in early-onset pre-eclampsia. A total of 120 patients and an equal number of women with normal pregnancy, from Government Maternity Hospital, Petlaburz, Hyderabad, India, were considered for the present study. A standard amplification refractory mutation system-polymerase chain reaction (ARMS-PCR) was carried out for genotyping followed by agarose gel electrophoresis. Appropriate statistical methods were applied to test for the significance of the results. It was found that the IL-10 -819 C allele (P = 0·003) and -592 A (P = 0·005) allele frequencies increased significantly in patients compared to controls. No significant difference was found with regard to -1082 promoter polymorphism. Haplotype analysis of the IL-10 single nucleotide polymorphisms (SNPs) revealed a significant association with ACC haplotype with a twofold increased risk in patients compared to controls. The frequencies of two common IL-10 haplotypes (GCC and ATA) did not show any significant difference. Further, the diplotype analysis revealed five genotypes: -1082A with -819C (P = 0·0016); -1082G with -819C (P = 0·0018); -819C with -592C (P = 0·001); -1082A with -592C (P = 0·032); and -1082G with -592C (P = 0·005) associated with the disease. These findings support the concept of contribution of IL-10 gene polymorphisms in the pathogenesis of early-onset pre-eclampsia. PMID:24962617

  12. Interleukin 10 gene promoter polymorphisms in women with early-onset pre-eclampsia

    PubMed Central

    Sowmya, S; Sri Manjari, K; Ramaiah, A; Sunitha, T; Nallari, P; Jyothy, A; Venkateshwari, A

    2014-01-01

    Pre-eclampsia is one of the most serious disorders of human pregnancy and T helper type 1 (Th1)/Th2 imbalance plays a major role in its aetiology. The Th2 cytokine, interleukin (IL)-10, plays a significant role in the maintenance of pregnancy. The present study is aimed at understanding the role of IL-10 promoter polymorphisms (−1082 G/A; −592 A/C and −819 C/T) and their haplotypes in early-onset pre-eclampsia. A total of 120 patients and an equal number of women with normal pregnancy, from Government Maternity Hospital, Petlaburz, Hyderabad, India, were considered for the present study. A standard amplification refractory mutation system–polymerase chain reaction (ARMS–PCR) was carried out for genotyping followed by agarose gel electrophoresis. Appropriate statistical methods were applied to test for the significance of the results. It was found that the IL-10 −819 C allele (P = 0·003) and −592 A (P = 0·005) allele frequencies increased significantly in patients compared to controls. No significant difference was found with regard to −1082 promoter polymorphism. Haplotype analysis of the IL-10 single nucleotide polymorphisms (SNPs) revealed a significant association with ACC haplotype with a twofold increased risk in patients compared to controls. The frequencies of two common IL-10 haplotypes (GCC and ATA) did not show any significant difference. Further, the diplotype analysis revealed five genotypes: −1082A with −819C (P = 0·0016); −1082G with −819C (P = 0·0018); −819C with −592C (P = 0·001); −1082A with −592C (P = 0·032); and −1082G with −592C (P = 0·005) associated with the disease. These findings support the concept of contribution of IL-10 gene polymorphisms in the pathogenesis of early-onset pre-eclampsia. PMID:24962617

  13. Relationship Between Interleukin-10 Gene C-819T Polymorphism and Gastric Cancer Risk: Insights From a Meta-Analysis

    PubMed Central

    Cui, Xigang; Huang, Qingxian; Li, Xianglin; Liu, Fang; Wang, Dan; Yan, Dong; Wang, Bin; Yang, Chunhua; Mi, Jia; Tian, Geng

    2016-01-01

    Background As a pleiotropic cytokine, interleukin-10 (IL-10) plays a regulatory role in carcinogenesis and tumor growth. The aim of this meta-analysis was to assess the susceptibility of the IL-10 gene C-819T polymorphism to gastric cancer. Material/Methods Study identification and data extraction were independently completed by 2 authors. Odds ratios (OR) and 95% confidence intervals (95% CI) were calculated and summarized. Results In total, 11 articles including 1960 gastric cancer patients and 3705 controls were qualified. Overall analyses revealed a 13% reduced risk of gastric cancer conferred by the −819T allele relative to the −819C allele (OR=0.87; 95% CI: 0.77–0.97; P=0.016), without heterogeneity (I2=35.1%). In subgroup analyses, a significant difference was identified in East Asian populations (OR=0.85; 95% CI: 0.73–0.98; P=0.029, I2=43.6%), for gastric adenocarcinoma (OR=0.80; 95% CI: 0.66–0.96; P=0.017, I2=0.0%), and in population-based studies (OR=0.81; 95% CI: 0.70–0.93; P=0.003, I2=0.0%). The visual funnel plots and Egger’s tests suggested no evidence of publication bias. Conclusions Extending previous findings, we demonstrate a protective role of the IL-10 gene −819T allele in susceptibility to gastric cancer, and this role was more evident for gastric adenocarcinoma. PMID:27516059

  14. Irsogladine Maleate Prevents Colitis in Interleukin-10 Gene-Deficient Mice by Reducing Interleukin-12 and -23 Production.

    PubMed

    Nakagawa, Tomoo; Katsuno, Tatsuro; Noguchi, Yoshiko; Mandai, Yasushi; Yoshihama, Sayuri; Saito, Keiko; Maruoka, Daisuke; Matsumura, Tomoaki; Arai, Makoto; Yokosuka, Osamu

    2015-01-01

    Irsogladine maleate (2,4-diamino-6-[2,5-dichlorophenyl]-s-triazine maleate; IM), an anti-peptic ulcer drug, may have a protective effect on the gastrointestinal mucosa. This study investigated the effects of IM on spontaneous colitis in interleukin-10 gene-deficient (IL-10(-/-)) mice. Five-week-old IL-10(-/-) mice were fed a control diet or one containing 100 ppm of IM for 10 weeks. Colonic tissues were evaluated morphologically and histologically. J774A.1 murine monocyte/macrophage cells were incubated with IM after lipopolysaccharide stimulation. mRNA expression was assessed by quantitative polymerase chain reaction (PCR) and protein concentration by enzyme-linked immunosorbent assay (ELISA). Colonic length, weight, and histological scores clearly demonstrated that spontaneous colitis was prevented in IL-10(-/-) mice fed a diet containing IM compared with those fed control diet. Levels of tumor necrosis factor-alpha (TNF-α) (-2.5-fold), IL-1β (-5.4), interferon-gamma (IFN-γ) (-4.5), IL-17 (-113.0), IL-12p35 (-21.0), IL-12p40 (-3.4), and IL-23p19 (-4.2) mRNA expression were significantly decreased in the colonic tissues of IM-treated animals, suggesting that oral treatment with IM suppressed the T-helper (Th)1/Th17 immune response in the colonic mucosa. An in vitro study using monocyte/macrophage cells to clarify the pharmacological action of IM indicated that IL-12p40 and IL-23p19 mRNA expression levels were dose-dependently decreased by IM treatment. ELISA showed that IL-12p40 and IL-23 protein secretion were significantly decreased by IM in a dose-dependent manner. Oral treatment with IM prevented spontaneous colitis in IL-10(-/-) mice by suppressing the colonic mucosal Th1/Th17 immune response through inhibition of IL-12 and -23 production in monocyte/macrophage cells. PMID:26521820

  15. Interleukin-10 but not transforming growth factor-β1 gene expression is up-regulated by vitamin D treatment in multiple sclerosis patients.

    PubMed

    Farsani, Zeinab Shirvani; Behmanesh, Mehrdad; Sahraian, Mohammad Ali

    2015-03-15

    Multiple sclerosis (MS) is an inflammatory and autoimmune disease. Variety of different genetics and environmental factors are involved in MS pathology. The epidemiological studies demonstrated that vitamin D has immune and immunomodulating effects on MS disease. Therefore, this study aims to evaluate the effect of vitamin D treatment on the expression of interleukin-10 (IL-10) and transforming growth factor-β1 (TGF-β1) genes in MS patients. We found that, the expression level of IL-10 gene in treated patients was up-regulated 3.84 times more than before treatment, but the expression level of TGF-β1 was not affected by vitamin D treatment. Also, a significant relationship was observed between vitamin D level and EDSS in MS patients. Our results indicated that the increased level of serum vitamin D and IL-10 gene expression may be associated with the reduction of EDSS scores in MS patients. PMID:25680585

  16. Polymorphisms in the interleukin-10 gene cluster are possibly involved in the increased risk for major depressive disorder

    PubMed Central

    Traks, Tanel; Koido, Kati; Eller, Triin; Maron, Eduard; Kingo, Külli; Vasar, Veiko; Vasar, Eero; Kõks, Sulev

    2008-01-01

    Background Innate immune inflammatory response is suggested to have a role in the pathogenesis of major depressive disorder (MDD). Interleukin (IL)-10 family cytokines IL-10, IL-19, IL-20, and IL-24 are all implicated in the inflammatory processes and polymorphisms in respective genes have been associated with various immunopathological conditions. This study was carried out to investigate whether single-nucleotide polymorphisms (SNPs) in these genes are also associated with MDD. Methods Case-control association study was performed with seven SNPs from the IL10 gene cluster. 153 patients with MDD and 277 healthy control individuals were recruited. Results None of the selected SNPs were individually associated with MDD. The linkage disequilibrium (LD) analysis indicated the existence of two recombination sites in the IL10 gene cluster, thus confirming the formerly established LD pattern of this genomic region. This also created two haplotype blocks, both consisting of three SNPs. Additionally, the haplotype analysis detected a significantly higher frequency of block 2 (IL20 and IL24 genes) haplotype TGC in the patients group compared to healthy control individuals (P = 0.0097). Conclusion Our study established increased risk for MDD related to the IL20 and IL24 haplotype and suggests that cytokines may contribute to the pathogenesis of MDD. Since none of the block 2 SNPs were individually associated with MDD, it is possible that other polymorphisms linked to them contribute to the disease susceptibility. Future studies are needed to confirm the results and to find the possible functional explanation. PMID:19087313

  17. Interleukin 10 (IL10) and transforming growth factor β1 (TGFβ1) gene polymorphisms in persistent IgE-mediated cow's milk allergy

    PubMed Central

    Jacob, Cristina Miuki Abe; Pastorino, Antonio Carlos; Okay, Thelma Suely; Castro, Ana Paula BM; Gushken, Andrea Keiko F.; Watanabe, Letícia Aki; Frucchi, Vanessa CZ; de Oliveira, Léa Campos

    2013-01-01

    OBJECTIVES: The aim of this cross-sectional study was to evaluate whether interleukin 10 (IL10) and transforming growth factor β1 (TGFβ1) gene polymorphisms were associated with persistent IgE-mediated cow's milk allergy in 50 Brazilian children. The diagnostic criteria were anaphylaxis triggered by cow's milk or a positive double-blind, placebo-controlled food challenge. Tolerance was defined as the absence of a clinical response to a double-blind, placebo-controlled food challenge or cow's milk exposure. METHOD: The genomic DNA of the 50 patients and 224 healthy controls (HCs) was used to investigate five IL10 gene polymorphisms (-3575A/T, -2849A/G, -2763A/C, -1082G/A, -592C/A) and one TGFβ1 polymorphism (-509C/T). RESULTS: Among the five IL10 polymorphisms analyzed, homozygosis for the G allele at the -1082 position was significantly higher in the patients compared with the healthy controls (p = 0.027) and in the persistent cow's milk allergy group compared with the healthy controls (p = 0.001). CONCLUSIONS: Homozygosis for the G allele at the IL10 -1082G/A polymorphism is associated with the persistent form of cow's milk allergy. PMID:23917667

  18. Changes in composition of caecal microbiota associated with increased colon inflammation in interleukin-10 gene-deficient mice inoculated with Enterococcus species.

    PubMed

    Bassett, Shalome A; Young, Wayne; Barnett, Matthew P G; Cookson, Adrian L; McNabb, Warren C; Roy, Nicole C

    2015-03-01

    Human inflammatory bowel disease (IBD) is a chronic intestinal disease where the resident microbiota contributes to disease development, yet the specific mechanisms remain unclear. Interleukin-10 gene-deficient (Il10-/-) mice develop inflammation similar to IBD, due in part to an inappropriate response to commensal bacteria. We have previously reported changes in intestinal morphology and colonic gene expression in Il10-/- mice in response to oral bacterial inoculation. In this study, we aimed to identify specific changes in the caecal microbiota associated with colonic inflammation in these mice. The microbiota was evaluated using pyrotag sequencing, denaturing gradient gel electrophoresis (DGGE) and quantitative real-time PCR. Microbiota profiles were influenced by genotype of the mice and by bacterial inoculation, and a strong correlation was observed between the microbiota and colonic inflammation scores. Although un-inoculated Il10-/- and C57 mice had similar microbiota communities, bacterial inoculation resulted in different changes to the microbiota in Il10-/- and C57 mice. Inoculated Il10-/- mice had significantly less total bacteria than un-inoculated Il10-/- mice, with a strong negative correlation between total bacterial numbers, relative abundance of Escherichia/Shigella, microbiota diversity, and colonic inflammation score. Our results show a putative causative role for the microbiota in the development of IBD, with potentially key roles for Akkermansia, or for Bacteroides, Helicobacter, Parabacteroides, and Alistipes, depending on the composition of the bacterial inoculum. These data support the use of bacterially-inoculated Il10-/- mice as an appropriate model to investigate human IBD. PMID:25768951

  19. Changes in Composition of Caecal Microbiota Associated with Increased Colon Inflammation in Interleukin-10 Gene-Deficient Mice Inoculated with Enterococcus Species

    PubMed Central

    Bassett, Shalome A.; Young, Wayne; Barnett, Matthew P. G.; Cookson, Adrian L.; McNabb, Warren C.; Roy, Nicole C.

    2015-01-01

    Human inflammatory bowel disease (IBD) is a chronic intestinal disease where the resident microbiota contributes to disease development, yet the specific mechanisms remain unclear. Interleukin-10 gene-deficient (Il10-/-) mice develop inflammation similar to IBD, due in part to an inappropriate response to commensal bacteria. We have previously reported changes in intestinal morphology and colonic gene expression in Il10-/- mice in response to oral bacterial inoculation. In this study, we aimed to identify specific changes in the caecal microbiota associated with colonic inflammation in these mice. The microbiota was evaluated using pyrotag sequencing, denaturing gradient gel electrophoresis (DGGE) and quantitative real-time PCR. Microbiota profiles were influenced by genotype of the mice and by bacterial inoculation, and a strong correlation was observed between the microbiota and colonic inflammation scores. Although un-inoculated Il10-/- and C57 mice had similar microbiota communities, bacterial inoculation resulted in different changes to the microbiota in Il10-/- and C57 mice. Inoculated Il10-/- mice had significantly less total bacteria than un-inoculated Il10-/- mice, with a strong negative correlation between total bacterial numbers, relative abundance of Escherichia/Shigella, microbiota diversity, and colonic inflammation score. Our results show a putative causative role for the microbiota in the development of IBD, with potentially key roles for Akkermansia, or for Bacteroides, Helicobacter, Parabacteroides, and Alistipes, depending on the composition of the bacterial inoculum. These data support the use of bacterially-inoculated Il10-/- mice as an appropriate model to investigate human IBD. PMID:25768951

  20. The association of three promoter polymorphisms in interleukin-10 gene with the risk for colorectal cancer and hepatocellular carcinoma: A meta-analysis

    PubMed Central

    Shi, Yan-Hui; Zhao, Dong-Mei; Wang, Yue-Fei; Li, Xue; Ji, Man-Ru; Jiang, Dan-Na; Xu, Bai-Ping; Zhou, Li; Lu, Chang-Zhu; Wang, Bin

    2016-01-01

    Mounting evidence supports a potent inhibitory role of interleukin-10 (IL-10) in tumor carcinogenesis, angiogenesis and metastasis. This meta-analysis was designed to examine the association of three promoter polymorphisms (−592C > A, −819C > T and −1082G > A) in IL-10 gene with the risk for colorectal cancer and hepatocellular carcinoma. Qualification assessment and data collection were completed by two authors independently. The random-effects model using the DerSimonian and Laird method was fitted by the STATA software. Twenty-five articles involving 5933 cases and 9724 controls were meta-analyzed. Overall comparisons of the mutant alleles (−592A, −819T and −1082A) of three promoter polymorphisms with alternative wild alleles failed to reveal any statistical significance for both colorectal cancer and hepatocellular carcinoma (P > 0.05), and the likelihood of heterogeneity was low (I2 < 50%). For −592C > A polymorphism, a significant risk for colorectal cancer was identified when analysis was restricted to East Asians (odds ratio or OR = 1.41, 95% confidence interval or CI: 1.18–1.68, P < 0.001) and retrospective studies (OR = 1.23, 95% CI: 1.09–1.39, P = 0.001). As weighed by the Egger’s test and the fill-and-trim method, there was a low probability of publication bias for all studied polymorphisms. Our findings collectively suggest that the −592C > A polymorphism in IL-10 gene might be a susceptibility locus for colorectal cancer in East Asians. PMID:27489033

  1. Interleukin-10 Alters Effector Functions of Multiple Genes Induced by Borrelia burgdorferi in Macrophages To Regulate Lyme Disease Inflammation ▿ †

    PubMed Central

    Gautam, Aarti; Dixit, Saurabh; Philipp, Mario T.; Singh, Shree R.; Morici, Lisa A.; Kaushal, Deepak; Dennis, Vida A.

    2011-01-01

    Interleukin-10 (IL-10) modulates inflammatory responses elicited in vitro and in vivo by Borrelia burgdorferi, the Lyme disease spirochete. How IL-10 modulates these inflammatory responses still remains elusive. We hypothesize that IL-10 inhibits effector functions of multiple genes induced by B. burgdorferi in macrophages to control concomitantly elicited inflammation. Because macrophages are essential in the initiation of inflammation, we used mouse J774 macrophages and live B. burgdorferi spirochetes as the model target cell and stimulant, respectively. First, we employed transcriptome profiling to identify genes that were induced by stimulation of cells with live spirochetes and that were perturbed by addition of IL-10 to spirochete cultures. Spirochetes significantly induced upregulation of 347 genes at both the 4-h and 24-h time points. IL-10 inhibited the expression levels, respectively, of 53 and 65 of the 4-h and 24-h genes, and potentiated, respectively, at 4 h and 24 h, 65 and 50 genes. Prominent among the novel identified IL-10-inhibited genes also validated by quantitative real-time PCR (qRT-PCR) were Toll-like receptor 1 (TLR1), TLR2, IRAK3, TRAF1, IRG1, PTGS2, MMP9, IFI44, IFIT1, and CD40. Proteome analysis using a multiplex enzyme-linked immunosorbent assay (ELISA) revealed the IL-10 modulation/and or potentiation of RANTES/CCL5, macrophage inflammatory protein 2 (MIP-2)/CXCL2, IP-10/CXCL10, MIP-1α/CCL3, granulocyte colony-stimulating factor (G-CSF)/CSF3, CXCL1, CXCL5, CCL2, CCL4, IL-6, tumor necrosis factor alpha (TNF-α), IL-1α, IL-1β, gamma interferon (IFN-γ), and IL-9. Similar results were obtained using sonicated spirochetes or lipoprotein as stimulants. Our data show that IL-10 alters effectors induced by B. burgdorferi in macrophages to control concomitantly elicited inflammatory responses. Moreover, for the first time, this study provides global insight into potential mechanisms used by IL-10 to control Lyme disease inflammation. PMID

  2. Protective role of adenovirus vector-mediated interleukin-10 gene therapy on endogenous islet β-cells in recent-onset type 1 diabetes in NOD mice

    PubMed Central

    LI, CHENG; ZHANG, LIJUAN; CHEN, YANYAN; LIN, XIAOJIE; LI, TANG

    2016-01-01

    The aim of the present study was to provide an animal experimental basis for the protective effect of the adenoviral vector-mediated interleukin-10 (Ad-mIL-10) gene on islet β-cells during the early stages of type 1 diabetes (T1D) in non-obese diabetic (NOD) mice. A total of 24 female NOD mice at the onset of diabetes were allocated at random into three groups (n=8 per group): Group 1, intraperitoneally injected with 0.1 ml Ad-mIL-10; group 2, intraperitoneally injected with 0.1 ml adenovirus vector; and group 3, was a diabetic control. In addition to groups 1, 2 and 3, 8 age- and gender-matched NOD mice were intraperitoneally injected with 0.1 ml PBS and assigned to group 4 as a normal control. All mice were examined weekly for body weight, urine glucose and blood glucose values prior to onset of diabetes, and at 1, 2 and 3 weeks after that, and all mice were sacrificed 3 weeks after injection. Serum levels of interleukin (IL)-10, interferon (IFN)-γ, IL-4, insulin and C-peptide were evaluated, and in addition the degree of insulitis and the local expression of IL-10 gene in the pancreas were detected. The apoptosis rate of pancreatic β-cells was determined using a TUNEL assay. Compared with groups 2 and 3, IL-10 levels in the serum and pancreas were elevated in group 1. Serum IFN-γ levels were decreased while serum IL-4 levels and IFN-γ/IL-4 ratio were significantly increased in group 1 (P<0.01). C-peptide and insulin levels were higher in group 1 compared with groups 2 and 3, (P<0.01). Furthermore, compared with groups 2 and 3, the degree of insulitis, islet β-cell apoptosis rate and blood glucose values did not change significantly (P>0.05). The administration of the Ad-mIL-10 gene induced limited immune regulatory and protective effects on islet β-cell function in NOD mice with early T1D, while no significant reduction in insulitis, islet β-cell apoptosis rate and blood glucose was observed. PMID:27168782

  3. Inflammatory Bowel Disease and Mutations Affecting the Interleukin-10 Receptor

    PubMed Central

    Glocker, Erik-Oliver; Kotlarz, Daniel; Boztug, Kaan; Gertz, E. Michael; Schäffer, Alejandro A.; Noyan, Fatih; Perro, Mario; Diestelhorst, Jana; Allroth, Anna; Murugan, Dhaarini; Hätscher, Nadine; Pfeifer, Dietmar; Sykora, Karl-Walter; Sauer, Martin; Kreipe, Hans; Lacher, Martin; Nustede, Rainer; Woellner, Cristina; Baumann, Ulrich; Salzer, Ulrich; Koletzko, Sibylle; Shah, Neil; Segal, Anthony W.; Sauerbrey, Axel; Buderus, Stephan; Snapper, Scott B.; Grimbacher, Bodo; Klein, Christoph

    2009-01-01

    BACKGROUND The molecular cause of inflammatory bowel disease is largely unknown. METHODS We performed genetic-linkage analysis and candidate-gene sequencing on samples from two unrelated consanguineous families with children who were affected by early-onset inflammatory bowel disease. We screened six additional patients with early-onset colitis for mutations in two candidate genes and carried out functional assays in patients’ peripheral-blood mononuclear cells. We performed an allogeneic hematopoietic stem-cell transplantation in one patient. RESULTS In four of nine patients with early-onset colitis, we identified three distinct homozygous mutations in genes IL10RA and IL10RB, encoding the IL10R1 and IL10R2 proteins, respectively, which form a heterotetramer to make up the interleukin-10 receptor. The mutations abrogate interleukin-10–induced signaling, as shown by deficient STAT3 (signal transducer and activator of transcription 3) phosphorylation on stimulation with interleukin-10. Consistent with this observation was the increased secretion of tumor necrosis factor α and other proinflammatory cytokines from peripheral-blood mononuclear cells from patients who were deficient in IL10R subunit proteins, suggesting that interleukin-10–dependent “negative feedback” regulation is disrupted in these cells. The allogeneic stem-cell transplantation performed in one patient was successful. CONCLUSIONS Mutations in genes encoding the IL10R subunit proteins were found in patients with early-onset enterocolitis, involving hyperinflammatory immune responses in the intestine. Allogeneic stem-cell transplantation resulted in disease remission in one patient. PMID:19890111

  4. Disease-Regulated Gene Therapy with Anti-Inflammatory Interleukin-10 Under the Control of the CXCL10 Promoter for the Treatment of Rheumatoid Arthritis.

    PubMed

    Broeren, Mathijs G A; de Vries, Marieke; Bennink, Miranda B; Arntz, Onno J; Blom, Arjen B; Koenders, Marije I; van Lent, Peter L E M; van der Kraan, Peter M; van den Berg, Wim B; van de Loo, Fons A J

    2016-03-01

    Disease-inducible promoters for the treatment of rheumatoid arthritis (RA) have the potential to provide regulated expression of therapeutic proteins in arthritic joints. In this study, we set out to identify promoters of human genes that are upregulated during RA and are suitable to drive the expression of relevant amounts of anti-inflammatory interleukin (IL)-10. Microarray analysis of RA synovial biopsies compared with healthy controls yielded a list of 22 genes upregulated during RA. Of these genes, CXCL10 showed the highest induction in lipopolysaccharide-stimulated synovial cells. The CXCL10 promoter was obtained from human cDNA and cloned into a lentiviral vector carrying firefly luciferase to determine the promoter inducibility in primary synovial cells and in THP-1 cells. The promoter activation was strongest 8-12 hr after stimulation with the proinflammatory cytokine tumor necrosis factor (TNF)-α and was reinducible after 96 hr. In addition, the CXCL10 promoter showed a significant response to RA patient serum, compared with sera from healthy individuals. The luciferase gene was replaced with IL-10 to determine the therapeutic properties of the CXCL10p-IL10 lentiviral vector. Primary synovial cells transduced with CXCL10p-IL10 showed a great increase in IL-10 production after stimulation, which reduced the release of proinflammatory cytokines TNF-α and IL-1β. We conclude that the selected proximal promoter of the CXCL10 gene responds to inflammatory mediators present in the serum of patients with RA and that transduction with the lentiviral CXCL10p-IL10 vector reduces inflammatory cytokine production by primary synovial cells from patients with RA. CXCL10 promoter-regulated IL-10 overexpression can thus provide disease-inducible local gene therapy suitable for RA. PMID:26711533

  5. Lactobacillus rhamnosus GG and Streptococcus thermophilus induce suppressor of cytokine signalling 3 (SOCS3) gene expression directly and indirectly via interleukin-10 in human primary macrophages

    PubMed Central

    Latvala, S; Miettinen, M; Kekkonen, R A; Korpela, R; Julkunen, I

    2011-01-01

    In the present study we have characterized T helper type 2 (Th2) [interleukin (IL)-10]/Th1 (IL-12) cytokine expression balance in human primary macrophages stimulated with multiple non-pathogenic Gram-positive bacteria used in the food industry and as probiotic substances. Bacteria representing Lactobacillus, Bifidobacterium, Lactococcus, Leuconostoc, Propionibacterium and Streptococcus species induced anti-inflammatory IL-10 production, although quantitative differences between the bacteria were observed. S. thermophilus was able to induce IL-12 production, while the production of IL-12 induced by other bacteria remained at a low level. The highest anti-inflammatory potential was seen with bifidobacteria, as evidenced by high IL-10/IL-12 induction ratios. All studied non-pathogenic bacteria were able to stimulate the expression of suppressor of cytokine signalling (SOCS) 3 that controls the expression of proinflammatory cytokine genes. Lactobacillus and Streptococcus species induced SOCS3 mRNA expression directly in the absence of protein synthesis and indirectly via bacteria-induced IL-10 production, as demonstrated by experiments with cycloheximide (CHX) and anti-IL-10 antibodies, respectively. The mitogen-activated protein kinase (MAPK) p38 signalling pathway played a key role in bacteria-induced SOCS3 gene expression. Enhanced IL-10 production and SOCS3 gene expression induced by live non-pathogenic Lactobacillus and Streptococcus is also likely to contribute to their immunoregulatory effects in vivo. PMID:21545585

  6. Interleukin-10 gene polymorphisms are associated with Behcet’s disease but not with Vogt-Koyanagi-Harada syndrome in the Chinese Han population

    PubMed Central

    Hu, Jianmin; Hou, Shengping; Zhu, Xueping; Fang, Jing; Zhou, Yan; Liu, Yunjia; Bai, Lin; Kijlstra, Aize

    2015-01-01

    Purpose This study aimed to investigate the association of interleukin (IL)-10 gene polymorphisms with Behcet’s disease (BD) and Vogt–Koyanagi–Harada (VKH) syndrome in the Chinese Han population. Methods A two-stage association study was performed on 718 BD patients, 300 VKH patients, and 1,753 controls. Genotyping of the IL-10 gene was performed for six single nucleotide polymorphisms (SNPs), including rs1800871, rs1800872, rs1800896, rs3021094, rs3790622, and rs1554286 using PCR-restricted fragment length polymorphism or TaqMan SNP assays. Real-time PCR was performed to test the IL-10 mRNA expression of the associated polymorphisms. Results The first-stage result showed significantly increased frequencies of the rs1800871 T allele, rs1800872 A allele, and rs1554286 T allele in BD patients compared with controls (Pcorrected (Pcorr) = 1.82×10−5, OR = 1.837; Pcorr = 6.1×10-5, OR = 1.780; Pcorr = 3.15×10−5, OR = 1.794, respectively). There was no association of the tested six SNPs with VKH syndrome. A second-stage study was therefore performed in BD patients to validate the result of the first stage, showing a significantly increased frequency of the rs1800871 T allele (Second stage, Pcorr = 5.59×10−5, OR = 1.493; Combined data, Pcorr = 3.65×10−11, OR = 1.632). Compared to the controls, an increased frequency of the rs1800871 T allele was observed in BD patients with extraocular findings, including genital ulcers, skin lesions, and a positive pathergy test. No difference was found among the mRNA expressions of IL-10 in the peripheral blood mononuclear cells (PBMCs) of controls with different genotypes of rs1800871 after stimulation of lipopolysaccharide (LPS) or anti-CD3/CD28 antibodies. Conclusions The findings showed that IL-10 is a risk gene for BD but not for VKH syndrome. PMID:26015771

  7. Interleukin 10 (- 1082 G/A) and (- 819 C/T) gene polymorphisms in Egyptian women with polycystic ovary syndrome (PCOS).

    PubMed

    Talaat, Roba M; Mohamed, Yasmin A; Mohamad, Ehab H; Elsharkawy, Marwa; Guirgis, Adel A

    2016-09-01

    Cytokines play critical roles in the pathogenesis of Polycystic Ovarian Syndrome (PCOS). This work was designed to study the implication of IL10 gene polymorphisms (- 1082 G/A and - 819 C/T) on the susceptibility of Egyptian women to have PCOS. Rotterdam consensus criteria were used to diagnose PCOS patients. Genotyping was performed by single-stranded polymorphism-polymerase chain reaction (SSP-PCR) in 61 PCOS patients and 80 healthy controls, and IL-10 serum levels were measured using Enzyme linked immunosorbent assay (ELISA). The frequency of IL10 - 1082 G/G (46%) genotype was significantly increased (p < 0.001) while the frequency of - 1082 A/A (16%) genotype was significantly decreased (p < 0.05) in PCOS patients compared to controls (14% and 35% for G/G and A/A genotypes; respectively). G allele (65%) is significantly increased (p < 0.01( in PCOS patients while A allele (61%) is significantly increased (p < 0.001( in control subjects. The distribution of IL10 -819 T/T genotype was significantly increased (p < 0.05) in PCOS group. G/G genotype (odd ratio (OR = 5.322) with confidence interval (CI = 2.364-11.982) and the G allele (OR = 2.828 with CI = 1.73-4.61) of - 1082 G/A and T/T genotype of - 819 C/T (OR = 4.18 with CI = 1.26-13.86) could be considered as risk factors for PCOS. IL-10 levels were significantly lower among PCOS patients (313.42 ± 30.10) compared to normal controls (4914.36 ± 303.72). Depending on our preliminary work, IL10 - 1082 G/G might be considered as a host genetic factor for PCOS susceptibility in Egyptian women. Studies concerning other cytokine gene polymorphisms are required to get a better understanding of the pathogenesis of PCOS disease. PMID:27617227

  8. Interleukin-10 and related cytokines and receptors.

    PubMed

    Pestka, Sidney; Krause, Christopher D; Sarkar, Devanand; Walter, Mark R; Shi, Yufang; Fisher, Paul B

    2004-01-01

    The Class 2 alpha-helical cytokines consist of interleukin-10 (IL-10), IL-19, IL-20, IL-22, IL-24 (Mda-7), and IL-26, interferons (IFN-alpha, -beta, -epsilon, -kappa, -omega, -delta, -tau, and -gamma) and interferon-like molecules (limitin, IL-28A, IL-28B, and IL-29). The interaction of these cytokines with their specific receptor molecules initiates a broad and varied array of signals that induce cellular antiviral states, modulate inflammatory responses, inhibit or stimulate cell growth, produce or inhibit apoptosis, and affect many immune mechanisms. The information derived from crystal structures and molecular evolution has led to progress in the analysis of the molecular mechanisms initiating their biological activities. These cytokines have significant roles in a variety of pathophysiological processes as well as in regulation of the immune system. Further investigation of these critical intercellular signaling molecules will provide important information to enable these proteins to be used more extensively in therapy for a variety of diseases. PMID:15032600

  9. A network map of Interleukin-10 signaling pathway.

    PubMed

    Verma, Renu; Balakrishnan, Lavanya; Sharma, Kusum; Khan, Aafaque Ahmad; Advani, Jayshree; Gowda, Harsha; Tripathy, Srikanth Prasad; Suar, Mrutyunjay; Pandey, Akhilesh; Gandotra, Sheetal; Prasad, T S Keshava; Shankar, Subramanian

    2016-03-01

    Interleukin-10 (IL-10) is an anti-inflammatory cytokine with important immunoregulatory functions. It is primarily secreted by antigen-presenting cells such as activated T-cells, monocytes, B-cells and macrophages. In biologically functional form, it exists as a homodimer that binds to tetrameric heterodimer IL-10 receptor and induces downstream signaling. IL-10 is associated with survival, proliferation and anti-apoptotic activities of various cancers such as Burkitt lymphoma, non-Hodgkins lymphoma and non-small scell lung cancer. In addition, it plays a central role in survival and persistence of intracellular pathogens such as Leishmania donovani, Mycobacterium tuberculosis and Trypanosoma cruzi inside the host. The signaling mechanisms of IL-10 cytokine are not well explored and a well annotated pathway map has been lacking. To this end, we developed a pathway resource by manually annotating the IL-10 induced signaling molecules derived from literature. The reactions were categorized under molecular associations, activation/inhibition, catalysis, transport and gene regulation. In all, 37 molecules and 76 reactions were annotated. The IL-10 signaling pathway can be freely accessed through NetPath, a resource of signal transduction pathways previously developed by our group. PMID:26253919

  10. High cerebrospinal fluid levels of interleukin-10 attained by AAV in dogs.

    PubMed

    Pleticha, J; Malkmus, S A; Heilmann, L F; Veesart, S L; Rezek, R; Xu, Q; Yaksh, T L; Beutler, A S

    2015-02-01

    Intrathecal (IT) gene transfer using adeno-associated virus (AAV) may be clinically promising as a treatment for chronic pain if it can produce sufficiently high levels of a transgene product in the cerebrospinal fluid (CSF). Although this strategy was developed in rodents, no studies investigating CSF levels of an analgesic or antiallodynic protein delivered by IT AAV have been performed in large animals. Interleukin-10 (IL-10) is an antiallodynic cytokine for which target therapeutic levels have been established in rats. The present study tested IT AAV8 encoding either human IL-10 (hIL-10) or enhanced green fluorescent protein (EGFP) in a dog model of IT drug delivery. AAV8/hIL-10 at a dose of 3.5 × 10(12) genome copies induced high hIL-10 levels in the CSF, exceeding the target concentration previously found to be antiallodynic in rodents by >1000-fold. AAV8/EGFP targeted the primary sensory and motor neurons and the meninges. hIL-10, a xenogeneic protein in dogs, induced anti-hIL-10 antibodies detectable in the CSF and serum of dogs. The high hIL-10 levels demonstrate the efficacy of AAV for delivery of secreted transgenes into the IT space of large animals, suggesting a strong case for further development toward clinical testing. PMID:25354684

  11. Interleukin-10.rs1800896 and Interleukin-18.rs1946518 gene polymorphisms could not predict the outcome of hepatitis C virus infection in Egyptian patients treated with pegylated interferon plus ribavirin.

    PubMed

    Abdelraheem, Wedad M; Hassuna, Noha A; Abuloyoun, Sahar M; Abdel Ghany, Hend M; Rizk, Hazem A; Abdelwahab, Sayed F

    2016-09-01

    A single-nucleotide polymorphism (SNP) in the interleukin (IL)-28B gene was used as a major predictor of the response to treatment in patients with hepatitis C virus (HCV) infection. Data examining the role of IL-10 and IL-18 gene polymorphisms among HCV genotype 4 (G4)-infected Egyptians in response to pegylated interferon (PEG-IFN) plus ribavirin (RBV) therapy are limited. This study investigated the impact of SNP at IL-10.rs1800896 (at position -1082) and IL-18.rs1946518 genes (at position -607) on the response to PEG-IFN/RBV therapy in HCV-infected Egyptians. This study was carried out on 100 HCV patients treated with PEG-IFN plus RBV and 100 healthy controls. The HCV patients included 50 treatment non-responders (NR) and 50 subjects with sustained virologic response (SVR). Genomic DNA from venous blood of subjects was extracted and IL-10.rs1800896 and IL-18.rs1946518 genotypes were determined using allele-specific amplification and SYBR Green real-time PCR. Linkage disequilibrium between the two SNPs was estimated using Haploview software. The frequency of the IL-10.rs1800896 AA, AG and GG genotypes among non-responders were 16 %, 70 % and 14 % while among SVR subjects, the frequency was 34 %, 60 % and 6 %, respectively (p=0.073). On the other hand, the frequency of the IL-18.rs1946518 AA, AC and CC genotypes among non-responders was 14 %, 50 % and 36 %, respectively, while among responders, these frequencies were 28 %, 44 % and 28 %, (p = 0.220). Both markers were in linkage equilibrium (D' = 0.23; r (2) = 0.052). SNPs in the IL-10.rs1800896 and IL-18.rs1946518 genes could not predict the outcome of HCV infection in Egyptians treated with PEG-IFN/RBV. PMID:27352267

  12. Role of cytokine gene (interferon-γ, transforming growth factor-β1, tumor necrosis factor-α, interleukin-6, and interleukin-10) polymorphisms in the risk of oral precancerous lesions in Taiwanese.

    PubMed

    Hsu, Han-Jen; Yang, Yi-Hsin; Shieh, Tien-Yu; Chen, Chung-Ho; Kao, Yu-Hsun; Yang, Chia-Fu; Ko, Edward Cheng-Chuan

    2014-11-01

    Oral squamous cell carcinoma can be preceded by some benign oral lesions with malignant potential, including leukoplakia, erythroplakia, oral lichen planus, and oral submucous fibrosis. There are different degrees of inflammatory cells infiltration in histopathology. Inflammatory cytokines may play a pathogenic role in the development of oral precancerous lesions (OPCLs). Genetic polymorphisms of cytokine-encoding genes are known to predispose to malignant disease. We hypothesized that the risk of OPCLs might be associated with cytokine gene polymorphisms of interferon (IFN)-γ, transforming growth factor (TGF)-β1, tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-10. In the present study, 42 OPCL patients and 128 controls were analyzed for eight polymorphisms in five different cytokine genes [IFN-γ (+874 T/A), TGF-β1 (codons 10 T/C and 25 G/C), TNF-α (-308 G/A), IL-6 (-174 G/C), and IL-10 (-1082 A/G, -819 T/C, and -592 A/C)]. Cytokine genotyping was determined by the polymerase chain reaction sequence-specific primer technique using commercial primers. Allele and genotype data were analyzed for significance of differences between cases and controls using the Chi-square (χ(2)) test. Two-sided p < 0.05 were considered to be statistically significant. A series of multivariate logistic regression models, adjusted for age, sex, betel quid chewing, alcohol consumption, and smoking, was constructed in order to access the contribution of homozygous or heterozygous variant genotypes of polymorphisms. The TNF-α (-308) polymorphism was significantly associated with OPCLs. There were significant differences in the distribution of AA, GA, and GG genotypes between OPCL patients and controls (p = 0.0004). Patients with the AA or GA genotype had a 3.63-fold increased risk of OPCLs. The TGF-β1 (codon 10 and 25) polymorphism was also significantly associated with OPCLs (p < 0.001). The IL-6 polymorphism was significantly associated with OPCLs. There are significant

  13. Interleukin-10 Family and Tuberculosis: An Old Story Renewed

    PubMed Central

    Abdalla, Abualgasim Elgaili; Lambert, Nzungize; Duan, Xiangke; Xie, Jianping

    2016-01-01

    The interleukin-10 (IL-10) family of cytokines consists of six immune mediators, namely IL-10, IL-19, IL-20, IL-22, IL-24 and IL-26. IL-10, IL-22, IL-24 and IL-26 are critical for the regulation of host defense against Mycobacterium tuberculosis infections. Specifically, IL-10 and IL-26 can suppress the antimycobacterial immunity and promote the survival of pathogen, while IL-22 and IL-24 can generate protective responses and inhibit the intracellular growth of pathogen. Knowledge about the new players in tuberculosis immunology, namely IL-10 family, can inform novel immunity-based countermeasures and host directed therapies against tuberculosis. PMID:27194948

  14. Interleukin-10 Family and Tuberculosis: An Old Story Renewed.

    PubMed

    Abdalla, Abualgasim Elgaili; Lambert, Nzungize; Duan, Xiangke; Xie, Jianping

    2016-01-01

    The interleukin-10 (IL-10) family of cytokines consists of six immune mediators, namely IL-10, IL-19, IL-20, IL-22, IL-24 and IL-26. IL-10, IL-22, IL-24 and IL-26 are critical for the regulation of host defense against Mycobacterium tuberculosis infections. Specifically, IL-10 and IL-26 can suppress the antimycobacterial immunity and promote the survival of pathogen, while IL-22 and IL-24 can generate protective responses and inhibit the intracellular growth of pathogen. Knowledge about the new players in tuberculosis immunology, namely IL-10 family, can inform novel immunity-based countermeasures and host directed therapies against tuberculosis. PMID:27194948

  15. Interleukin-10 Gene Therapy-Mediated Amelioration of Bacterial Pneumonia

    PubMed Central

    Morrison, Daniel F.; Foss, Dennis L.; Murtaugh, Michael P.

    2000-01-01

    Respiratory infection by Actinobacillus pleuropneumoniae causes a highly pathogenic necrotizing pleuropneumonia with severe edema, hemorrhage and fever. Acute infection is characterized by expression of inflammatory cytokines, including interleukin-1 (IL-1), IL-6 and IL-8. To determine if high level production of inflammatory cytokines contributed to disease pathogenesis, we investigated if inhibiting macrophage activation with adenovirus type 5-expressed IL-10 (Ad-5/IL-10) reduced the severity of acute disease. Porcine tracheal epithelial cells infected with Ad-5/IL-10 produced bioactive human IL-10. When pigs were intratracheally infected with A. pleuropneumoniae, pigs pretreated with Ad-5/IL-10 showed a significant reduction in the amount of lung damage when compared to adenovirus type 5-expressing β-galactosidase (Ad-5/β-Gal)-treated and untreated pigs. In addition, serum zinc levels were unchanged, the lung weight/body weight ratio (an indicator of vascular leakage) was significantly reduced, and lung pathology scores were reduced. Myeloperoxidase activity in lung lavage fluid samples, an indicator of neutrophil invasion, was decreased to levels similar to that seen in pigs not infected with A. pleuropneumoniae. Reduction in inflammatory cytokine levels in lung lavage fluid samples correlated with the clinical observations in that pigs pretreated with Ad-5/IL-10 showed a corresponding reduction of IL-1 and tumor necrosis factor (TNF) compared with untreated and Ad-5/β-Gal-treated pigs. IL-6 levels were unaffected by pretreatment with Ad-5/IL-10, consistent with observations that IL-6 was not derived from alveolar macrophages. Since inflammatory cytokines are expressed at high levels in acute bacterial pleuropneumonia, these results indicate that macrophage activation, involving overproduction of IL-1 and TNF, is a prime factor in infection-related cases of massive lung injury. PMID:10899882

  16. Family-based association study of interleukin 10 (IL10) and interleukin 10 receptor alpha (IL10RA) functional polymorphisms in schizophrenia in Polish population.

    PubMed

    Kapelski, Pawel; Skibinska, Maria; Maciukiewicz, Malgorzata; Pawlak, Joanna; Zaremba, Dorota; Twarowska-Hauser, Joanna

    2016-08-15

    Schizophrenia is a heterogeneous disorder and its etiology remains incompletely elucidated. Among possible causes, immunological factors have been implicated in its pathogenesis and course. Interleukin-10 (IL10) and it's receptor IL10RA may play an important role for immunological aspects in etiologies of major psychiatric disorders including schizophrenia. The aim of this study was to perform a transmission disequilibrium test (TDT) on a group of 146 schizophrenia trios from the Polish population. Functional polymorphisms from IL10 (rs1800872, rs1800871, rs1800896, rs1800890, and rs6676671) and IL10RA (rs3135932 and rs2229113) genes were analyzed. A lack of association with schizophrenia was detected for IL10 and IL10RA single polymorphisms and haplotypes. PMID:27397081

  17. Lateral gene transfer in eukaryotes.

    PubMed

    Andersson, J O

    2005-06-01

    Lateral gene transfer -- the transfer of genetic material between species -- has been acknowledged as a major mechanism in prokaryotic genome evolution for some time. Recently accumulating data indicate that the process also occurs in the evolution of eukaryotic genomes. However, there are large rate variations between groups of eukaryotes; animals and fungi seem to be largely unaffected, with a few exceptions, while lateral gene transfer frequently occurs in protists with phagotrophic lifestyles, possibly with rates comparable to prokaryotic organisms. Gene transfers often facilitate the acquisition of functions encoded in prokaryotic genomes by eukaryotic organisms, which may enable them to colonize new environments. Transfers between eukaryotes also occur, mainly into larger phagotrophic eukaryotes that ingest eukaryotic cells, but also between plant lineages. These findings have implications for eukaryotic genomic research in general, and studies of the origin and phylogeny of eukaryotes in particular. PMID:15761667

  18. Interleukin-10 overexpression in macrophages suppresses atherosclerosis in hyperlipidemic mice

    PubMed Central

    Han, Xinbing; Kitamoto, Shiro; Wang, Hongwei; Boisvert, William A.

    2010-01-01

    In atherogenesis, macrophage foam cell formation is modulated by pathways involving both the uptake and efflux of cholesterol. We recently showed that interleukin-10 (IL-10) modulates lipid metabolism by enhancing both uptake and efflux of cholesterol in macrophages. However, the mechanistic details of these properties in vivo have been unclear. Thus, the purpose of this study was to determine whether expression of IL-10 in macrophages would alter susceptibility to atherosclerosis and whether IL-10 exerts its antiatherosclerotic properties by modulating lipid metabolism in macrophages. We utilized a macrophage-specific retroviral vector that allows long-term in vivo expression of IL-10 in macrophages through transplantation of retrovirally transduced bone marrow cells (BMCs). IL-10 expressed by macrophages derived from transduced BMCs inhibited atherosclerosis in LDLR−/− mice by reducing cholesteryl ester accumulation in atherosclerotic sites. Experiments with primary macrophages indicated that macrophage source of IL-10 stimulated both the uptake (by up-regulating scavenger receptors) and efflux of cholesterol (by activating the PPARγ-LXR-ABCA1/ABCG1 pathway), thereby reducing inflammation and apoptosis in atherosclerosis. These findings indicate that BMC-transduced macrophage IL-10 production can act as a strong antiatherogenic agent, and they highlight a novel antiatherosclerotic therapy using a simple, yet effective, stem cell transduction system that facilitates long-term expression of IL-10 in macrophages.—Han, X., Kitamoto, S., Wang, H., Boisvert, W. A. Interleukin-10 overexpression in macrophages suppresses atherosclerosis in hyperlipidemic mice. PMID:20354139

  19. Gene Transfer into Cardiac Myocytes

    PubMed Central

    Lang, Sarah E.; Westfall, Margaret V.

    2016-01-01

    Traditional methods for DNA transfection are often inefficient and toxic for terminally differentiated cells, such as cardiac myocytes. Vector-based gene transfer is an efficient approach for introducing exogenous cDNA into these types of primary cell cultures. In this chapter, separate protocols for adult rat cardiac myocyte isolation and gene transfer with recombinant adenovirus are provided and are routinely utilized for studying the effects of sarcomeric proteins on myofilament function. PMID:25836585

  20. Significant roles played by interleukin-10 in outcome of pregnancy

    PubMed Central

    Mobini, Masoud; Mortazavi, Maryam; Nadi, Somayeh; Zare-Bidaki, Mohammad; Pourtalebi, Somayeh; Arababadi, Mohammad Kazemi

    2016-01-01

    Imbalanced immune responses against fetus alloantigens can lead to abnormality in pregnancy. Interleukin-10 (IL-10) plays key roles in regulation of immune responses against self and foreign antigens to induce tolerance to these antigens. Therefore, alteration in expression of IL-10 during pregnancy may result in several pathologic conditions such as preterm labor. IL-10 leads to a normal pregnancy via several molecular mechanisms including development of tolerogenic dendritic cells, T regulatory lymphocytes and activation of the JAK1/STAT3 pathway in the target cells. This review has collected recent data regarding the status of IL-10 expression during term and preterm deliveries and also its molecular mechanisms that lead to a normal pregnancy. PMID:27081455

  1. Gene transfer for erythropoiesis enhancement.

    PubMed

    Naffakh, N; Danos, O

    1996-08-01

    The spectrum of anemias treated with recombinant human erythropoietin is rapidly broadening. Lifelong treatment with very high doses is now under evaluation for beta-thalassemia and sickle cell anemia. These indications make it worthwhile to search for methods that will allow a permanent systemic delivery of the hormone. Here, we review experimental gene-transfer-based procedures for erythropoietin delivery in vivo. In mice, both ex vivo and direct in vivo approaches for gene transfer have resulted in the long-term production of therapeutic levels of the hormone. Gene transfer of erythropoietin could become a viable alternative to the injection of the purified recombinant protein once reliable procedures for controlling transgene expression are available. PMID:8796920

  2. Gene transfer: transduction.

    PubMed

    Frangipani, Emanuela

    2014-01-01

    Bacteriophages able to propagate on Pseudomonas strains are very common and can be easily isolated from natural environments or lysogenic strains. The development of transducing systems has allowed bacterial geneticists to perform chromosome analyses and mutation mapping. Moreover, these systems have also been proved to be a successful tool for molecular microbiologists to introduce a foreign gene or a mutation into the chromosome of a bacterial cell. This chapter provides a description of the phage methodology illustrated by Adams in 1959 and applicable to strain PAO1 derivatives. PMID:24818891

  3. Role of Interleukin 10 Transcriptional Regulation in Inflammation and Autoimmune Disease

    PubMed Central

    Iyer, Shankar Subramanian; Cheng, Genhong

    2012-01-01

    Interleukin 10 (IL-10) is a cytokine with potent anti-inflammatory properties that plays a central role in limiting host immune response to pathogens, thereby preventing damage to the host and maintaining normal tissue homeostasis. Dysregulation of IL-10 is associated with enhanced immunopathology in response to infection as well as increased risk for development of many autoimmune diseases. Thus a fundamental understanding of IL-10 gene expression is critical for our comprehension of disease progression and resolution of host inflammatory response. In this review, we discuss modes of regulation of IL-10 gene expression in immune effector cell types, including signal transduction, epigenetics, promoter architecture, and post-transcriptional regulation, and how aberrant regulation contributes to immunopathology and disease progression. PMID:22428854

  4. Gene transfer in intact animals

    NASA Astrophysics Data System (ADS)

    Cline, M. J.; Stang, H.; Mercola, K.; Morse, L.; Ruprecht, R.; Browne, J.; Salser, W.

    1980-04-01

    Resistance to methotrexate was induced in bone marrow cells of mice by transformation in vitro with DNA from a drug-resistant cell line. Transformed cells were injected in vivo and haematopoietic cells expressing resistance were selected by drug treatment of recipients. Transformed cells had elevated levels of dihydrofolate reductase and demonstrated a proliferative advantage over untransformed cells, indicating successful gene transfer.

  5. Horizontal gene transfer in plants.

    PubMed

    Gao, Caihua; Ren, Xiaodong; Mason, Annaliese S; Liu, Honglei; Xiao, Meili; Li, Jiana; Fu, Donghui

    2014-03-01

    Horizontal gene transfer (HGT) describes the transmission of genetic material across species boundaries. HGT often occurs in microbic and eukaryotic genomes. However, the pathways by which HGTs occur in multicellular eukaryotes, especially in plants, are not well understood. We systematically summarized more than ten possible pathways for HGT. The intimate contact which frequently occurs in parasitism, symbiosis, pathogen, epiphyte, entophyte, and grafting interactions could promote HGTs between two species. Besides these direct transfer methods, genes can be exchanged with a vector as a bridge: possible vectors include pollen, fungi, bacteria, viruses, viroids, plasmids, transposons, and insects. HGT, especially when involving horizontal transfer of transposable elements, is recognized as a significant force propelling genomic variation and biological innovation, playing an important functional and evolutionary role in both eukaryotic and prokaryotic genomes. We proposed possible mechanisms by which HGTs can occur, which is useful in understanding the genetic information exchange among distant species or distant cellular components. PMID:24132513

  6. Production of interleukin-10 by human bronchogenic carcinoma.

    PubMed Central

    Smith, D. R.; Kunkel, S. L.; Burdick, M. D.; Wilke, C. A.; Orringer, M. B.; Whyte, R. I.; Strieter, R. M.

    1994-01-01

    Interleukin-10 (IL-10) is a recently characterized cytokine with suppressive activity against various aspects of the cellular immune response. Our laboratory has previously demonstrated that another anti-inflammatory cytokine, IL-1 receptor antagonist (IRAP) is produced and secreted by human bronchogenic carcinomas. We speculated that tumor production of IRAP may mitigate host responses and confer increased tumor viability. In this study, we investigated the capacity of human bronchogenic tumors to produce IL-10 as another possible mechanism to attenuate host defenses. We found increased levels of antigenic IL-10 in tissue homogenates of human bronchogenic carcinomas compared with normal lung tissue (13.69 +/- 2.87 versus 5.84 +/- 0.84 ng/mg total protein). Immunohistochemical staining of tumors illustrate primary localization of antigenic IL-10 to individual tumor cells. Analysis of supernatants of several unstimulated human bronchogenic cell lines in vitro demonstrated the ability of tumor cells to constitutively produce IL-10. Functional studies of mononuclear cells, cultured in the presence of conditioned medium from a bronchogenic cell line, demonstrated their increased tumor necrosis factor and IL-6 production with the addition of neutralizing antibodies to IL-10. These findings demonstrate that human bronchogenic carcinomas elaborate functional IL-10, which may significantly impair immune effector cell function and enable the tumor to evade host defenses. Images Figure 1 Figure 2 PMID:8030748

  7. Matrine ameliorates spontaneously developed colitis in interleukin-10-deficient mice.

    PubMed

    Wu, Cong; Xu, Zheng; Gai, Renhua; Huang, Kehe

    2016-07-01

    Interleukin-10 (IL-10)-deficient mice spontaneously develop T cell-mediated colitis. Previous reports have shown that Matrine may reduce the symptoms of acute colitis induced by trinitrobenzene sulfonic acid (TNBS). However, whether Matrine impacts chronic colitis remains unknown. In this study, we investigated whether Matrine could limit the symptoms of spontaneously developed colitis and its potential molecular mechanisms. IL-10 deficient mice were given Matrine or a PBS control by oral gavage daily for 4weeks and were euthanized at week 2 or week 4. We measured body weight, colon length and weight, and histological scores. We also evaluated the spontaneous secretion of IL-12/23p40, IFN-γ and IL-17 in colon explant cultures as well as IFN-γ and IL-17 secretion in unseparated mesenteric lymph node (MLN) cells, and assessed IFN-γ, IL-17, IL-1β and IL-6 mRNA expression in colon tissue. In addition, we analyzed the proportions of CD4-positive and CD8-positive cells in unseparated MLN cells. Our results show that Matrine-treated mice exhibited better body weight recovery than controls and that histological scores and spontaneously secreted IL-12/23p40, IFN-γ and IL-17 in colon tissue were significantly decreased in treated mice compared with controls. The proportion of CD4-positive cells of MLNs in treated mice was significantly smaller than that in controls at week 4. Both cytokine production and mRNA expression of IFN-γ and IL-17 were significantly reduced in treated mice compared with controls. Taken together, our results indicate that Matrine may ameliorate spontaneously developed chronic colitis and could be considered as a therapeutic alternative for chronic colitis. PMID:27179305

  8. Effects of interleukin-10 on cutaneous wounds and scars in humans of African continental ancestral origin.

    PubMed

    Kieran, Ingrid; Taylor, Catherine; Bush, Jim; Rance, Mark; So, Karen; Boanas, Adam; Metcalfe, Anthony; Hobson, Rosalind; Goldspink, Nick; Hutchison, John; Ferguson, Mark

    2014-01-01

    Scars in humans of African continental ancestry heal with an exaggerated inflammatory response and a generally wider scar. Interleukin-10 is an anti-inflammatory and antifibrotic cytokine. A randomized controlled trial in Caucasians found that exogenous interleukin-10 resulted in improved macroscopic scar appearance and reduced scar redness. We investigated the effects of interleukin-10 on cutaneous scarring in volunteers of African ancestral origin in an exploratory, single-center, within-subject, double-blind randomized controlled trial. Fifty-six subjects received two of four potential prerandomized concentrations of interleukin-10 (5, 25, 100, and 250 ng/100 µL) in two full-thickness incisions on the upper inner arms. Anatomically matching incisions on the contralateral arm were treated with placebo. Scars were excised at 1 month for histological analysis and were redosed with the same regimen. Resultant excision scars were followed up for 12 months for scar width measurement and scoring. Scoring was performed by trial doctors, subjects, and a panel. Incisions treated with 100 ng/100 µL interleukin-10 had significantly reduced microscopic scar widths. Incisions treated with 5 and 25 ng/100 µL interleukin-10 were also narrower, but not significantly. There were no differences observed in pro-inflammatory or pro-fibrotic markers between interleukin-10 and placebo treatment. There was no long-term evidence that 100 ng/100 µL interleukin-10 had a therapeutic effect on macroscopic scar width or appearance, as excisions treated with this concentration were significantly wider than placebo between 8 and 12 months of maturation. Doctors showed a trend toward favoring the macroscopic appearance of placebo-treated excisions compared with those treated with 250 ng/100 µL interleukin-10. Panelists scored placebo-treated excisions as significantly better-appearing than those treated with 250 ng/100 µL interleukin-10. Doctors' scores showed a

  9. Protective role of interleukin-10 in Ozone-induced pulmonary inflammation**

    EPA Science Inventory

    Background: The mechanisms underlying ozone (03)-induced pulmonary inflammation remain unclear. Interleukin-10 (IL-10) is an anti-inflammatory cytokine that is known to inhibit inflammatory mediators. Objectives: We investigated the molecular mechanisms underlying interleuken-10...

  10. Interleukin 10 suppresses experimental chronic, granulomatous inflammation induced by bacterial cell wall polymers.

    PubMed Central

    Herfarth, H H; Mohanty, S P; Rath, H C; Tonkonogy, S; Sartor, R B

    1996-01-01

    BACKGROUND AND AIMS: Interleukin 10 (IL10) inhibits monocyte/macrophage and T lymphocyte effector functions. This study examined the effect of systemically administered IL10 on acute and chronic granulomatous enterocolitis, hepatitis, and arthritis in a rat model. METHODS: Lewis rats were injected intramurally with streptococcal peptidoglycan-polysaccharide (PG-APS) polymers. Beginning 12 hours before PG-APS injection, rats were treated daily with subcutaneous murine recombinant IL10 or vehicle for three or 17 days. RESULTS: IL10 attenuated acute enterocolitis in a dose dependent fashion (p < 0.01). Protective effects were more profound in the chronic granulomatous phase with decreased enterocolitis and markedly inhibited leucocytosis, hepatic granulomas, and chronic erosive arthritis (p < 0.001). IL10 downregulated tissue IL1, IL6, tumour necrosis factor alpha, and interferon gamma gene expression, consistent with the in vitro effects of IL10 on PG-APS-stimulated splenocytes. Caecal IL1 protein concentrations and IL2 and interferon gamma secretion by in vitro stimulated mesenteric lymph nodes were downregulated in IL10 treated animals. CONCLUSIONS: These results indicate that exogenous IL10 can inhibit experimental granulomatous inflammatory responses and suggest that IL10 treatment could be an effective new therapeutic approach in human disorders such as Crohn's disease, rheumatoid arthritis, and sarcoidosis. Images Figure 1 Figure 3 Figure 5 Figure 6 PMID:9038666

  11. Lateral Gene Transfer from the Dead

    PubMed Central

    Szöllősi, Gergely J.; Tannier, Eric; Lartillot, Nicolas; Daubin, Vincent

    2013-01-01

    In phylogenetic studies, the evolution of molecular sequences is assumed to have taken place along the phylogeny traced by the ancestors of extant species. In the presence of lateral gene transfer, however, this may not be the case, because the species lineage from which a gene was transferred may have gone extinct or not have been sampled. Because it is not feasible to specify or reconstruct the complete phylogeny of all species, we must describe the evolution of genes outside the represented phylogeny by modeling the speciation dynamics that gave rise to the complete phylogeny. We demonstrate that if the number of sampled species is small compared with the total number of existing species, the overwhelming majority of gene transfers involve speciation to and evolution along extinct or unsampled lineages. We show that the evolution of genes along extinct or unsampled lineages can to good approximation be treated as those of independently evolving lineages described by a few global parameters. Using this result, we derive an algorithm to calculate the probability of a gene tree and recover the maximum-likelihood reconciliation given the phylogeny of the sampled species. Examining 473 near-universal gene families from 36 cyanobacteria, we find that nearly a third of transfer events (28%) appear to have topological signatures of evolution along extinct species, but only approximately 6% of transfers trace their ancestry to before the common ancestor of the sampled cyanobacteria. [Gene tree reconciliation; lateral gene transfer; macroevolution; phylogeny.] PMID:23355531

  12. A Single Amino Acid Determines the Immunostimulatory Activity of Interleukin 10

    PubMed Central

    Ding, Yaozhong; Qin, Lihui; Kotenko, Serguei V.; Pestka, Sidney; Bromberg, Jonathan S.

    2000-01-01

    Cellular interleukin 10s (cIL-10s) of human and murine origin have extensive sequence and structural homology to the Epstein-Barr virus BCRF-I gene product, known as viral IL-10 (vIL-10). Although these cytokines share many immunosuppressive properties, vIL-10 lacks several of the immunostimulatory activities of cIL-10 on certain cell types. The molecular and cellular bases for this dichotomy are not currently defined. Here, we show that the single amino acid isoleucine at position 87 of cIL-10 is required for its immunostimulatory function. Substitution of isoleucine in cIL-10 with alanine, which corresponds to the vIL-10 residue, abrogates immunostimulatory activity for thymocytes, mast cells, and alloantigenic responses while preserving immunosuppressive activity for inhibition of interferon γ production and prolongation of cardiac allograft survival. Conversely, substitution of alanine with isoleucine in vIL-10 converts it to a cIL-10–like molecule with immunostimulatory activity. This single conservative residue alteration significantly affects ligand affinity for receptor; however, affinity changes do not necessarily alter specific activities for biologic responses in a predictable fashion. These results suggest complex regulation of IL-10 receptor–ligand interactions and subsequent biological responses. These results demonstrate that vIL-10 may represent a captured and selectively mutated cIL-10 gene that benefits viral pathogenesis by leading to ineffective host immune responses. The ability to manipulate the activity of IL-10 in either a stimulatory or suppressive direction may be of practical value for regulating immune responses for disease therapy, and of theoretical value for determining what aspects of IL-10 activity are important for normal T cell responses. PMID:10637267

  13. Horizontal gene transfer between bacteria and animals.

    PubMed

    Dunning Hotopp, Julie C

    2011-04-01

    Horizontal gene transfer is increasingly described between bacteria and animals. Such transfers that are vertically inherited have the potential to influence the evolution of animals. One classic example is the transfer of DNA from mitochondria and chloroplasts to the nucleus after the acquisition of these organelles by eukaryotes. Even today, many of the described instances of bacteria-to-animal transfer occur as part of intimate relationships such as those of endosymbionts and their invertebrate hosts, particularly insects and nematodes, while numerous transfers are also found in asexual animals. Both of these observations are consistent with modern evolutionary theory, in particular the serial endosymbiotic theory and Muller's ratchet. Although it is tempting to suggest that these particular lifestyles promote horizontal gene transfer, it is difficult to ascertain given the nonrandom sampling of animal genome sequencing projects and the lack of a systematic analysis of animal genomes for such transfers. PMID:21334091

  14. A Multihit Model: Colitis Lessons from the Interleukin-10–deficient Mouse

    PubMed Central

    Keubler, Lydia M.; Buettner, Manuela; Häger, Christine

    2015-01-01

    Abstract: Complex mechanisms are pulling the strings to initiate the development of inflammatory bowel disease. Current evidence indicates that an interaction of genetic susceptibilities (polymorphisms), environmental factors, and the host microbiota leads to a dysregulation of the mucosal immune system. In the past decades, the interleukin-10–deficient mouse has served as an excellent model to mirror the multifactorial nature of this disease. Here, we want to review in detail the interplay of the genetic factors, immune aspects, and especially summarize and discuss the role of the microbiota contributing to colitis development in the interleukin-10–deficient mouse model of inflammatory bowel disease as a multihit model. PMID:26164667

  15. High expression hampers horizontal gene transfer.

    PubMed

    Park, Chungoo; Zhang, Jianzhi

    2012-01-01

    Horizontal gene transfer (HGT), the movement of genetic material from one species to another, is a common phenomenon in prokaryotic evolution. Although the rate of HGT is known to vary among genes, our understanding of the cause of this variation, currently summarized by two rules, is far from complete. The first rule states that informational genes, which are involved in DNA replication, transcription, and translation, have lower transferabilities than operational genes. The second rule asserts that protein interactivity negatively impacts gene transferability. Here, we hypothesize that high expression hampers HGT, because the fitness cost of an HGT to the recipient, arising from the 1) energy expenditure in transcription and translation, 2) cytotoxic protein misfolding, 3) reduction in cellular translational efficiency, 4) detrimental protein misinteraction, and 5) disturbance of the optimal protein concentration or cell physiology, increases with the expression level of the transferred gene. To test this hypothesis, we examined laboratory and natural HGTs to Escherichia coli. We observed lower transferabilities of more highly expressed genes, even after controlling the confounding factors from the two established rules and the genic GC content. Furthermore, expression level predicts gene transferability better than all other factors examined. We also confirmed the significant negative impact of gene expression on the rate of HGTs to 127 of 133 genomes of eubacteria and archaebacteria. Together, these findings establish the gene expression level as a major determinant of horizontal gene transferability. They also suggest that most successful HGTs are initially slightly deleterious, fixed because of their negligibly low costs rather than high benefits to the recipient. PMID:22436996

  16. Horizontal gene transfer, genome innovation and evolution.

    PubMed

    Gogarten, J Peter; Townsend, Jeffrey P

    2005-09-01

    To what extent is the tree of life the best representation of the evolutionary history of microorganisms? Recent work has shown that, among sets of prokaryotic genomes in which most homologous genes show extremely low sequence divergence, gene content can vary enormously, implying that those genes that are variably present or absent are frequently horizontally transferred. Traditionally, successful horizontal gene transfer was assumed to provide a selective advantage to either the host or the gene itself, but could horizontally transferred genes be neutral or nearly neutral? We suggest that for many prokaryotes, the boundaries between species are fuzzy, and therefore the principles of population genetics must be broadened so that they can be applied to higher taxonomic categories. PMID:16138096

  17. A Causal Role of Genetically Elevated Circulating Interleukin-10 in the Development of Digestive Cancers

    PubMed Central

    Niu, Wenquan; Pang, Qing; Lin, Ting; Wang, Zhixin; Zhang, Jingyao; Tai, Minghui; Zhang, Lingqiang; Zhang, Li; Gu, Mingliang; Liu, Chang; Qu, Kai

    2016-01-01

    Abstract Recent studies have observed a high level of circulating interleukin-10 (IL-10) in patients with digestive cancers, yet whether elevated IL-10 is causally associated with digestive cancers so far remained unresolved. We therefore meta-analyzed available observational studies with Mendelian randomization method to explore this causal association by employing IL-10 gene 3 variants (-592C>A, -819C>T, and -1082A>G) as instruments. Data were available from 52 articles encompassing 29,307 subjects. Subgroup analysis by cancer type indicated that -1082A>G was associated with increased risk of gastric cancer (odds ratio [OR] = 1.19; 95% confidence interval [CI]: 1.05–1.35; P = 0.006), and the association was reinforced for intestinal type gastric cancer (OR = 1.26; 95%CI: 1.09–1.44; P = 0.001). By ethnicity, risk estimate for -1082G allele carriers was increased by 21% for digestive cancers in East Asians (95%CI: 1.05–1.40; P = 0.009). As for the genotype–phenotype association, carriers of -1082G allele had an overall 20.21 pg/mL higher IL-10 level than those with -1082AA genotype (P = 0.023). In further Mendelian randomization analysis, the predicted OR for 10 pg/mL increment in IL-10 was 1.14 (95%CI: 1.01–16.99) in gastric cancer. Our findings provided evidence for a causal role of genetically elevated IL-10 in the development of gastric cancer, especially in East Asians and for intestinal type gastric cancer. PMID:26886630

  18. Biliverdin Reductase A (BVRA) Mediates Macrophage Expression of Interleukin-10 in Injured Kidney

    PubMed Central

    Hu, Zhizhi; Pei, Guangchang; Wang, Pengge; Yang, Juan; Zhu, Fengmin; Guo, Yujiao; Wang, Meng; Yao, Ying; Zeng, Rui; Liao, Wenhui; Xu, Gang

    2015-01-01

    Biliverdin reductase A is an enzyme, with serine/threonine/tyrosine kinase activation, converting biliverdin (BV) to bilirubin (BR) in heme degradation pathway. It has been reported to have anti-inflammatory and antioxidant effect in monocytes and human glioblastoma. However, the function of BVRA in polarized macrophage was unknown. This study aimed to investigate the effect of BVRA on macrophage activation and polarization in injured renal microenvironment. Classically activated macrophages (M1macrophages) and alternative activation of macrophages (M2 macrophages) polarization of murine bone marrow derived macrophage was induced by GM-CSF and M-CSF. M1 polarization was associated with a significant down-regulation of BVRA and Interleukin-10 (IL-10), and increased secretion of TNF-α. We also found IL-10 expression was increased in BVRA over-expressed macrophages, while it decreased in BVRA knockdown macrophages. In contrast, BVRA over-expressed or knockdown macrophages had no effect on TNF-α expression level, indicating BVRA mediated IL-10 expression in macrophages. Furthermore, we observed in macrophages infected with recombinant adenoviruses BVRA gene, which BVRA over-expressed enhanced both INOS and ARG-1 mRNA expression, resulting in a specific macrophage phenotype. Through in vivo study, we found BVRA positive macrophages largely existed in mice renal ischemia perfusion injury. With the treatment of the regular cytokines GM-CSF, M-CSF or LPS, excreted in the injured renal microenvironment, IL-10 secretion was significantly increased in BVRA over-expressed macrophages. In conclusion, the BVRA positive macrophage is a source of anti-inflammatory cytokine IL-10 in injured kidney, which may provide a potential target for treatment of kidney disease. PMID:26393580

  19. Gene Transfers Between Distantly Related Organisms

    NASA Technical Reports Server (NTRS)

    Doolittle, Russell F.

    2003-01-01

    With the completion of numerous microbial genome sequences, reports of individual gene transfers between distantly related prokaryotes have become commonplace. On the other hand, transfers between prokaryotes and eukaryotes still excite the imagination. Many of these claims may be premature, but some are certainly valid. In this chapter, the kinds of supporting data needed to propose transfers between distantly related organisms and cite some interesting examples are considered.

  20. Horizontal gene transfer of stress resistance genes through plasmid transport.

    PubMed

    Shoeb, Erum; Badar, Uzma; Akhter, Jameela; Shams, Hina; Sultana, Maria; Ansari, Maqsood A

    2012-03-01

    The horizontal gene transfer of plasmid-determined stress tolerance was achieved under lab conditions. Bacterial isolates, Enterobacter cloacae (DGE50) and Escherichia coli (DGE57) were used throughout the study. Samples were collected from contaminated marine water and soil to isolate bacterial strains having tolerance against heavy metals and antimicrobial agents. We have demonstrated plasmid transfer, from Amp(+)Cu(+)Zn(-) strain (DGE50) to Amp(-)Cu(-)Zn(+) strain (DGE57), producing Amp(+)Cu(+)Zn(+) transconjugants (DGE(TC50→57)) and Amp(+)Cu(-)Zn(+) transformants (DGE(TF50→57)). DGE57 did not carry any plasmid, therefore, it can be speculated that zinc tolerance gene in DGE57 is located on chromosome. DGE50 was found to carry three plasmids, out of which two were transferred through conjugation into DGE57, and only one was transferred through transformation. Plasmid transferred through transformation was one out of the two transferred through conjugation. Through the results of transformation it was revealed that the genes of copper and ampicillin tolerance in DGE50 were located on separate plasmids, since only ampicillin tolerance genes were transferred through transformation as a result of one plasmid transfer. By showing transfer of plasmids under lab conditions and monitoring retention of respective phenotype via conjugation and transformation, it is very well demonstrated how multiple stress tolerant strains are generated in nature. PMID:22805823

  1. In vitro gene transfer by electrosonoporation.

    PubMed

    Escoffre, J M; Kaddur, K; Rols, M P; Bouakaz, A

    2010-10-01

    Among the nonviral methods for gene delivery in vitro, electroporation is simple, inexpensive and safe. To upregulate the expression level of transfected gene, we investigated the applicability of electrosonoporation. This approach consists of a combination of electric pulses and ultrasound assisted with gas microbubbles. Cells were first electroporated with plasmid DNA encoding-enhanced green fluorescent protein and then sonoporated in presence of contrast microbubbles. Twenty-four hours later, cells that received electrosonoporation demonstrated a four-fold increase in transfection level and a six-fold increase in transfection efficiency compared with cells having undergone electroporation alone. Although electroporation induced the formation of DNA aggregates into the cell membrane, sonoporation induced its direct propulsion into the cytoplasm. Sonoporation can improve the transfer of electro-induced DNA aggregates by allowing its free and rapid entrance into the cells. These results demonstrated that in vitro gene transfer by electrosonoporation could provide a new potent method for gene transfer. PMID:20850028

  2. Gene transfer mediated by alpha2-macroglobulin.

    PubMed Central

    Schneider, H; Huse, K; Birkenmeier, G; Otto, A; Scholz, G H

    1996-01-01

    alpha2-Macroglobulin covalently linked to poly(L)-lysine can be used as a vehicle for receptor-mediated gene transfer. This modified alpha2-macroglobulin maintains its ability to bind to the alpha2-macroglobulin receptor, and was shown to introduce a luciferase reporter gene plasmid into HepG2 human hepatoma cells in vitro. The alpha2-macroglobulin receptor is a very large and multifunctional cell surface receptor, whose rapid and efficient internalization rate makes it attractive for gene therapy, e.g. for hepatic gene targeting via injection into the portal vein. PMID:8871570

  3. Modulation of the inflammatory response by increasing fetal wound size or interleukin-10 overexpression determines wound phenotype and scar formation.

    PubMed

    Morris, Michael W; Allukian, Myron; Herdrich, Benjamin J; Caskey, Robert C; Zgheib, Carlos; Xu, Junwang; Dorsett-Martin, Wanda; Mitchell, Marc E; Liechty, Kenneth W

    2014-01-01

    Wound size impacts the threshold between scarless regeneration and reparative healing in the fetus with increased inflammation showed in fetal scar formation. We hypothesized that increased fetal wound size increases pro-inflammatory and fibrotic genes with resultant inflammation and fibroplasia and that transition to scar formation could be reversed by overexpression of interleukin-10 (IL-10). To test this hypothesis, 2-mm and 8-mm dermal wounds were created in mid-gestation fetal sheep. A subset of 8-mm wounds were injected with a lentiviral vector containing the IL-10 transgene (n = 4) or vehicle (n = 4). Wounds were harvested at 3 or 30 days for histology, immunohistochemistry, analysis of gene expression by microarray, and validation with real-time polymerase chain reaction. In contrast to the scarless 2-mm wounds, 8-mm wounds showed scar formation with a differential gene expression profile, increased inflammatory cytokines, decreased CD45+ cells, and subsequent inflammation. Lentiviral-mediated overexpression of the IL-10 gene resulted in conversion to a regenerative phenotype with decreased inflammatory cytokines and regeneration of dermal architecture. In conclusion, increased fetal wounds size leads to a unique gene expression profile that promotes inflammation and leads to scar formation and furthermore, these results show the significance of attenuated inflammation and IL-10 in the transition from fibroplasia to fetal regenerative healing. PMID:24844340

  4. Hypothermia increases interleukin-6 and interleukin-10 in juvenile endotoxemic mice

    PubMed Central

    Stewart, Corrine R.; Landseadel, Jessica P.; Gurka, Matthew J.; Fairchild, Karen D.

    2013-01-01

    Objective To develop a juvenile mouse model to establish effects of in vivo hypothermia on expression of the inflammation-modulating cytokines tumor necrosis factor-α, interleukin-1β, interleukin-6, and interleukin-10. Although induced hypothermia is neuroprotective in some patients, the mechanisms of protection are not well understood and concerns remain over potential detrimental effects, particularly in the setting of infection. We previously showed that in vitro hypothermia increases production of tumor necrosis factor-α and interleukin-1β in lipopolysaccharide-treated monocytes. Design Laboratory investigation. Setting Research laboratory. Subjects Juvenile (4-wk) male C57BL/6 mice. Interventions Mice were given chlorpromazine to suspend thermoregulation and lipopolysaccharide to stimulate cytokine production. Core temperature was maintained at 32°C or 37°C for 6 hrs by adjusting environmental temperature. In separate experiments, lipopolysaccharide-treated mice were kept in a cooling chamber without chlorpromazine treatment. Measurements and Main Results Plasma and organs were collected for cytokine quantitation. Chlorpromazine-treated hypothermic mice had 2.3-fold and 1.8-fold higher plasma interleukin-6 and interleukin-10 levels at 6 hrs compared with identically treated normothermic mice (p < .05), whereas plasma tumor necrosis factor-α and interleukin-1β were not significantly different at 2 hrs or 6 hrs. Liver tumor necrosis factor-α and interleukin-6 were significantly higher in hypothermic vs. normothermic mice, but lung and brain cytokines were not different. Lipopolysaccharide-treated mice kept in a cooling chamber without chlorpromazine treatment developed varying degrees of hypothermia with associated increases in plasma interleukin-6 and interleukin-10. A nonspecific marker of stress (plasma corticosterone) was not affected by hypothermia in lipopolysaccharide-treated mice. Conclusion Further studies are necessary to determine the

  5. Viral Vectors for in Vivo Gene Transfer

    NASA Astrophysics Data System (ADS)

    Thévenot, E.; Dufour, N.; Déglon, N.

    The transfer of DNA into the nucleus of a eukaryotic cell (gene transfer) is a central theme of modern biology. The transfer is said to be somatic when it refers to non-germline organs of a developed individual, and germline when it concerns gametes or the fertilised egg of an animal, with the aim of transmitting the relevant genetic modification to its descendents [1]. The efficient introduction of genetic material into a somatic or germline cell and the control of its expression over time have led to major advances in understanding how genes work in vivo, i.e., in living organisms (functional genomics), but also to the development of innovative therapeutic methods (gene therapy). The efficiency of gene transfer is conditioned by the vehicle used, called the vector. Desirable features for a vector are as follows: Easy to produce high titer stocks of the vector in a reproducible way. Absence of toxicity related to transduction (transfer of genetic material into the target cell, and its expression there) and no immune reaction of the organism against the vector and/or therapeutic protein. Stability in the expression of the relevant gene over time, and the possibility of regulation, e.g., to control expression of the therapeutic protein on the physiological level, or to end expression at the end of treatment. Transduction of quiescent cells should be as efficient as transduction of dividing cells. Vectors currently used fall into two categories: non-viral and viral vectors. In non-viral vectors, the DNA is complexed with polymers, lipids, or cationic detergents (described in Chap. 3). These vectors have a low risk of toxicity and immune reaction. However, they are less efficient in vivo than viral vectors when it comes to the number of cells transduced and long-term transgene expression. (Naked DNA transfer or electroporation is rather inefficient in the organism. This type of gene transfer will not be discussed here, and the interested reader is referred to the

  6. Protection against Paracoccidioides brasiliensis infection in mice treated with modulated dendritic cells relies on inhibition of interleukin-10 production by CD8+ T cells.

    PubMed

    da Costa, Thiago Alves; Di Gangi, Rosária; Martins, Paula; Longhini, Ana Leda Figueiredo; Zanucoli, Fábio; de Oliveira, Alexandre Leite Rodrigues; Stach-Machado, Dagmar Ruth; Burger, Eva; Verinaud, Liana; Thomé, Rodolfo

    2015-11-01

    Paracoccidioidomycosis is a systemic infection prevalent in Latin American countries. Disease develops after inhalation of Paracoccidioides brasiliensis conidia followed by an improper immune activation by the host leucocytes. Dendritic cells (DCs) are antigen-presenting cells with the unique ability to direct the adaptive immune response by the time of activation of naive T cells. This study was conducted to test whether extracts of P. brasiliensis would induce maturation of DCs. We found that DCs treated with extracts acquired an inflammatory phenotype and upon adoptive transfer conferred protection to infection. Interestingly, interleukin-10 production by CD8(+) T cells was ablated following DC transfer. Further analyses showed that lymphocytes from infected mice were high producers of interleukin-10, with CD8(+) T cells being the main source. Blockage of cross-presentation to CD8(+) T cells by modulated DCs abolished the protective effect of adoptive transfer. Collectively, our data show that adoptive transfer of P. brasiliensis-modulated DCs is an interesting approach for the control of infection in paracoccidioidomycosis. PMID:26302057

  7. Unsupervised learning in detection of gene transfer.

    PubMed

    Hamel, L; Nahar, N; Poptsova, M S; Zhaxybayeva, O; Gogarten, J P

    2008-01-01

    The tree representation as a model for organismal evolution has been in use since before Darwin. However, with the recent unprecedented access to biomolecular data, it has been discovered that, especially in the microbial world, individual genes making up the genome of an organism give rise to different and sometimes conflicting evolutionary tree topologies. This discovery calls into question the notion of a single evolutionary tree for an organism and gives rise to the notion of an evolutionary consensus tree based on the evolutionary patterns of the majority of genes in a genome embedded in a network of gene histories. Here, we discuss an approach to the analysis of genomic data of multiple genomes using bipartition spectral analysis and unsupervised learning. An interesting observation is that genes within genomes that have evolutionary tree topologies, which are in substantial conflict with the evolutionary consensus tree of an organism, point to possible horizontal gene transfer events which often delineate significant evolutionary events. PMID:18509479

  8. Herpes simplex virus vector-mediated expression of interleukin-10 reduces below-level central neuropathic pain after spinal cord injury

    PubMed Central

    Lau, Darryl; Harte, Steven E.; Morrow, Thomas J.; Wang, Shiyong; Mata, Marina; Fink, David J.

    2012-01-01

    Background Neuroimmune activation in the spinal dorsal horn plays an important role in the pathogenesis of chronic pain after peripheral nerve injury. Objective We wanted to examine the role of neuroimmune activation in below level neuropathic pain after traumatic spinal cord injury (SCI). Methods Right hemilateral SCI was created in male Sprague Dawley rats by controlled blunt impact through a T12 laminectomy. Pain related behaviors were assessed using both evoked reflex responses and an operant conflict-avoidance test. Neuroimmune activation was blocked by the anti-inflammatory cytokine interleukin-10 (IL-10) delivered by a non-replicating herpes simplex virus (HSV)-based gene transfer vector (vIL10). Markers of neuroimmune activation were assessed using immunohistochemistry and Western blot. Results One week after SCI, injured animals demonstrated mechanical allodynia, thermal hyperalgesia, and mechanical hyperalgesia in the hind limbs below the level of injury. Animals inoculated with vIL10 had a statistically significant reduction in all of these measures compared to injured rats or injured rats inoculated with control vector. Conflict-avoidance behavior of injured rats inoculated with vIL10 was consistent with significantly reduced pain compared to injured rats injected with control vector. These behavioral results correlated with a significant decrease in spinal tumor necrosis factor α (mTNFα) expression assessed by Western blot and astrocyte activation assessed by glial fibrillary acidic protein immunohistochemistry. Conclusion Below level pain after SCI is characterized by neuroimmune activation (increase mTNFα and astrocyte activation). Blunting of the neuroimmune response by HSV-mediated delivery of IL-10 reduced pain-related behaviors, and may represent a potential novel therapeutic agent. PMID:22593113

  9. Lateral gene transfer in the subsurface

    SciTech Connect

    Barkay, Tamar; Sobecky, Patricia

    2007-08-27

    Lateral gene transfer (LGT) is an important adaptive mechanism among prokaryotic organisms. This mechanism is particularly important for the response of microorganisms to changing environmental conditions because it facilitates the transfer of a large number of genes and their rapid expression. Together the transferred genes promote rapid genetic and metabolic changes that may enhance survival to newly established and sometimes hostile environmental conditions. The goal of our project was to examine if and how LGT enhances microbial adaptation to toxic heavy metals in subsurface environments that had been contaminated by mixed wastes due to activities associated with the production of nuclear energy and weapons. This task has been accomplished by dividing the project to several sub-tasks. Thus, we: (1) Determined the level of resistance of subsurface bacterial isolates to several toxic metals, all identified as pollutants of concern in subsurface environments; (2) Designed, tested, and applied, a molecular approach that determined whether metal resistance genes had evolved by LGT among subsurface bacteria; and (3) Developed a DNA hybridization array for the identification of broad host range plasmids and of metal resistance plasmids. The results are briefly summarized below with references to published papers and manuscripts in preparation where details about our research can be found. Additional information may be found in copies of our published manuscripts and conference proceedings, and our yearly reports that were submitted through the RIMS system.

  10. Clinical Applications Involving CNS Gene Transfer

    PubMed Central

    Kantor, Boris; McCown, Thomas; Leone, Paola; Gray, Steven J.

    2015-01-01

    Diseases of the central nervous system (CNS) have traditionally been the most difficult to treat by traditional pharmacological methods, due mostly to the blood–brain barrier and the difficulties associated with repeated drug administration targeting the CNS. Viral vector gene transfer represents a way to permanently provide a therapeutic protein within the nervous system after a single administration, whether this be a gene replacement strategy for an inherited disorder or a disease-modifying protein for a disease such as Parkinson's. Gene therapy approaches for CNS disorders has evolved considerably over the last two decades. Although a breakthrough treatment has remained elusive, current strategies are now considerably safer and potentially much more effective. This chapter will explore the past, current, and future status of CNS gene therapy, focusing on clinical trials utilizing adeno-associated virus and lentiviral vectors. PMID:25311921

  11. Perinatal Gene Transfer to the Liver

    PubMed Central

    McKay, Tristan R; Rahim, Ahad A; Buckley, Suzanne M.K; Ward, Natalie J; Chan, Jerry K.Y; Howe, Steven J; Waddington, Simon N

    2011-01-01

    The liver acts as a host to many functions hence raising the possibility that any one may be compromised by a single gene defect. Inherited or de novo mutations in these genes may result in relatively mild diseases or be so devastating that death within the first weeks or months of life is inevitable. Some diseases can be managed using conventional medicines whereas others are, as yet, untreatable. In this review we consider the application of early intervention gene therapy in neonatal and fetal preclinical studies. We appraise the tools of this technology, including lentivirus, adenovirus and adeno-associated virus (AAV)-based vectors. We highlight the application of these for a range of diseases including hemophilia, urea cycle disorders such as ornithine transcarbamylase deficiency, organic acidemias, lysosomal storage diseases including mucopolysaccharidoses, glycogen storage diseases and bile metabolism. We conclude by assessing the advantages and disadvantages associated with fetal and neonatal liver gene transfer. PMID:21774770

  12. Evaluation of Interleukin-10 Levels in Patients Diagnosed with Chronic Hepatitis

    PubMed Central

    Özgüler, M; Akbulut, HH; Akbulut, A

    2015-01-01

    ABSTRACT Objective: One of the most important factors playing a role in chronic hepatitis B pathogenesis is cytokine release and one of the cytokines with anti-inflammatory characteristic is interleukin-10 (IL-10). The aim of the present study is to examine IL-10 levels in patients with chronic hepatitis B. Subjects and Methods: Sixty-three patients with chronic hepatitis B disease who had not received any antiviral treatment were included in the study. Serum IL-10 level was investigated by enzyme-linked immunosorbent assay (ELISA) method. In the control group, 25 healthy individuals with mean age similar to the patient population were included. Control and patient groups were compared and data were statistically analysed. Results: Interleukin-10 levels of 25 patients with hepatitis B virus (HBV) DNA levels between 2000 and 20 000 IU/mL were compared with those of 25 subjects in the control group, and the level in the chronic hepatitis B group was statistically significantly higher (p < 0.05). Interleukin-10 levels of 38 patients with HBV DNA > 20 000 IU/mL were statistically significantly higher than those in the control group. When chronic hepatitis B patients were compared among themselves, IL-10 levels increased as HBV DNA levels increased. Also, when IL-10 levels of hepatitis B ‘e’ antigen (HBeAg) positive patients were compared with those of HBeAg negative patients, the difference was not statistically significant. Conclusion: It is believed that decreasing IL-10 levels by various methods would have significant contributions in disease progression and treatment. Moreover, IL-10 level may be an important marker in HBeAg seroconversion and evaluation of treatment response. PMID:26360676

  13. Foamy virus vectors for gene transfer

    PubMed Central

    Trobridge, Grant D.

    2009-01-01

    Foamy virus (FV) vectors are efficient gene delivery vehicles that have shown great promise for gene therapy in preclinical animal models. FVs or spumaretroviruses are not endemic in humans, but are prevalent in nonhuman primates and in other mammals. They have evolved means for efficient horizontal transmission in their host species without pathology. FV vectors have several unique properties that make them well-suited for therapeutic gene transfer including a desirable safety profile, a broad tropism, a large transgene capacity, and the ability to persist in quiescent cells. They mediate efficient and stable gene transfer to hematopoietic stem cells (HSCs) in mouse models, and in the canine large animal model. Analysis of FV vector integration sites in vitro and in hematopoietic repopulating cells shows they have a unique integration profile, and suggests they may be safer than gammaretroviruses or lentiviral vectors. Here properties of FVs relevant to the safety and efficacy of FV vectors are discussed. The development of FV vector systems is described, and studies evaluating their potential in vitro, and in small and large animal models is reviewed. PMID:19743892

  14. Horizontal Gene Transfer, Dispersal and Haloarchaeal Speciation

    PubMed Central

    Papke, R. Thane; Corral, Paulina; Ram-Mohan, Nikhil; de la Haba, Rafael R.; Sánchez-Porro, Cristina; Makkay, Andrea; Ventosa, Antonio

    2015-01-01

    The Halobacteria are a well-studied archaeal class and numerous investigations are showing how their diversity is distributed amongst genomes and geographic locations. Evidence indicates that recombination between species continuously facilitates the arrival of new genes, and within species, it is frequent enough to spread acquired genes amongst all individuals in the population. To create permanent independent diversity and generate new species, barriers to recombination are probably required. The data support an interpretation that rates of evolution (e.g., horizontal gene transfer and mutation) are faster at creating geographically localized variation than dispersal and invasion are at homogenizing genetic differences between locations. Therefore, we suggest that recurrent episodes of dispersal followed by variable periods of endemism break the homogenizing forces of intrapopulation recombination and that this process might be the principal stimulus leading to divergence and speciation in Halobacteria. PMID:25997110

  15. Interleukin-10 and tumour necrosis factor-alpha serum levels in chronic Chagas disease patients.

    PubMed

    Vasconcelos, R H T; Azevedo, E de A N; Diniz, G T N; Cavalcanti, M da G A de M; de Oliveira, W; de Morais, C N L; Gomes, Y de M

    2015-07-01

    In Chagas disease, chronically infected individuals may be asymptomatic or may present cardiac or digestive complications, and it is well known that the human immune response is related to different clinical manifestations. Different patterns of cytokine levels have been previously described in different clinical forms of this disease, but contradictory results are reported. Our aim was to evaluate the serum levels of interleukin-10 and tumour necrosis factor-alpha in patients with asymptomatic and cardiac Chagas disease. The serum interleukin-10 levels in patients with cardiomyopathy were higher than those in asymptomatic patients, mainly in those without heart enlargement. Although no significant difference was observed in serum tumour necrosis factor-alpha levels among the patients, we found that cardiac patients also present high levels of this cytokine, largely those with heart dilatation. Therefore, these cytokines play an important role in chronic Chagas disease cardiomyopathy. Follow-up investigations of these and other cytokines in patients with chronic Chagas disease need to be conducted to improve the understanding of the immunopathology of this disease. PMID:25728555

  16. Simple rapid method for gene transfer

    SciTech Connect

    Cockburn, A.F.; Meier, H.

    1990-01-30

    The object of the present invention is to provide methods for gene transfer that reduce or eliminate cellular pretreatment steps, e.g., the removal of cell wall by chemical or enzymatic methods, is rapid and can be practiced without the need of additional expensive equipment. Cells, embryos or tissues selected for genetic manipulation are suspended in an Eppendorf tube in an aliquot of the desired genetic material to be transferred to which the resulting mixture is added and is agitated by vortexing from about 30 to about 90 seconds. The cells, embryos or tissue are sedimented and the DNA supernatant removed. After sedimentation, the injected material is resuspended in or on a growth medium to assay for expression.

  17. Gene Transfer between Salmonella enterica Serovar Typhimurium inside Epithelial Cells

    PubMed Central

    Ferguson, Gayle C.; Heinemann, Jack A.; Kennedy, Martin A.

    2002-01-01

    Virulence and antibiotic resistance genes transfer between bacteria by bacterial conjugation. Conjugation also mediates gene transfer from bacteria to eukaryotic organisms, including yeast and human cells. Predicting when and where genes transfer by conjugation could enhance our understanding of the risks involved in the release of genetically modified organisms, including those being developed for use as vaccines. We report here that Salmonella enterica serovar Typhimurium conjugated inside cultured human cells. The DNA transfer from donor to recipient bacteria was proportional to the probability that the two types of bacteria occupied the same cell, which was dependent on viable and invasive bacteria and on plasmid tra genes. Based on the high frequencies of gene transfer between bacteria inside human cells, we suggest that such gene transfers occur in situ. The implications of gene transfer between bacteria inside human cells, particularly in the context of antibiotic resistance, are discussed. PMID:11914355

  18. Reducible cationic lipids for gene transfer.

    PubMed Central

    Wetzer, B; Byk, G; Frederic, M; Airiau, M; Blanche, F; Pitard, B; Scherman, D

    2001-01-01

    One of the main challenges of gene therapy remains the increase of gene delivery into eukaryotic cells. We tested whether intracellular DNA release, an essential step for gene transfer, could be facilitated by using reducible cationic DNA-delivery vectors. For this purpose, plasmid DNA was complexed with cationic lipids bearing a disulphide bond. This reduction-sensitive linker is expected to be reduced and cleaved in the reducing milieu of the cytoplasm, thus potentially improving DNA release and consequently transfection. The DNA--disulphide-lipid complexation was monitored by ethidium bromide exclusion, and the size of complexes was determined by dynamic light scattering. It was found that the reduction kinetics of disulphide groups in DNA--lipid complexes depended on the position of the disulphide linker within the lipid molecule. Furthermore, the internal structure of DNA--lipid particles was examined by small-angle X-ray scattering before and after lipid reduction. DNA release from lipid complexes was observed after the reduction of disulphide bonds of several lipids. Cell-transfection experiments suggested that complexes formed with selected reducible lipids resulted in up to 1000-fold higher reporter-gene activity, when compared with their analogues without disulphide bonds. In conclusion, reduction-sensitive groups introduced into cationic lipid backbones potentially allow enhanced DNA release from DNA--lipid complexes after intracellular reduction and represent a tool for improved vectorization. PMID:11389682

  19. Interleukin 10 Receptor Signaling: Master Regulator of Intestinal Mucosal Homeostasis in Mice and Humans

    PubMed Central

    Shouval, Dror S.; Ouahed, Jodie; Biswas, Amlan; Goettel, Jeremy A.; Horwitz, Bruce H.; Klein, Christoph; Muise, Aleixo M.; Snapper, Scott B.

    2016-01-01

    Interleukin 10 (IL10) is a key anti-inflammatory cytokine that can inhibit proinflammatory responses of both innate and adaptive immune cells. An association between IL10 and intestinal mucosal homeostasis became clear with the discovery that IL10 and IL10 receptor (IL10R)-deficient mice develop spontaneous intestinal inflammation. Similarly, patients with deleterious mutations in IL10, IL10RA, or IL10RB present with severe enterocolitis within the first months of life. Here, we review recent findings on how IL10- and IL10R-dependent signaling modulates innate and adaptive immune responses in the murine gastrointestinal tract, with implications of their role in the prevention of inflammatory bowel disease (IBD). In addition, we discuss the impact of IL10 and IL10R signaling defects in humans and their relationship to very early-onset IBD (VEO-IBD). PMID:24507158

  20. Horizontal gene transfer from Agrobacterium to plants

    PubMed Central

    Matveeva, Tatiana V.; Lutova, Ludmila A.

    2014-01-01

    Most genetic engineering of plants uses Agrobacterium mediated transformation to introduce novel gene content. In nature, insertion of T-DNA in the plant genome and its subsequent transfer via sexual reproduction has been shown in several species in the genera Nicotiana and Linaria. In these natural examples of horizontal gene transfer from Agrobacterium to plants, the T-DNA donor is assumed to be a mikimopine strain of A. rhizogenes. A sequence homologous to the T-DNA of the Ri plasmid of Agrobacterium rhizogenes was found in the genome of untransformed Nicotiana glauca about 30 years ago, and was named “cellular T-DNA” (cT-DNA). It represents an imperfect inverted repeat and contains homologs of several T-DNA oncogenes (NgrolB, NgrolC, NgORF13, NgORF14) and an opine synthesis gene (Ngmis). A similar cT-DNA has also been found in other species of the genus Nicotiana. These presumably ancient homologs of T-DNA genes are still expressed, indicating that they may play a role in the evolution of these plants. Recently T-DNA has been detected and characterized in Linaria vulgaris and L. dalmatica. In Linaria vulgaris the cT-DNA is present in two copies and organized as a tandem imperfect direct repeat, containing LvORF2, LvORF3, LvORF8, LvrolA, LvrolB, LvrolC, LvORF13, LvORF14, and the Lvmis genes. All L. vulgaris and L. dalmatica plants screened contained the same T-DNA oncogenes and the mis gene. Evidence suggests that there were several independent T-DNA integration events into the genomes of these plant genera. We speculate that ancient plants transformed by A. rhizogenes might have acquired a selective advantage in competition with the parental species. Thus, the events of T-DNA insertion in the plant genome might have affected their evolution, resulting in the creation of new plant species. In this review we focus on the structure and functions of cT-DNA in Linaria and Nicotiana and discuss their possible evolutionary role. PMID:25157257

  1. Interleukin 10 (Tenovil) in the prevention of postoperative recurrence of Crohn's disease

    PubMed Central

    Colombel, J; Rutgeerts, P; Malchow, H; Jacyna, M; Nielsen, O; Rask-Madsen, J; Van Deventer, S; Ferguson, A; Desreumaux, P; Forbes, A; Geboes, K; Melani, L; Cohard, M

    2001-01-01

    BACKGROUND AND AIMS—New lesions of Crohn's disease occur early after ileal or ileocolonic resection and ileocolonic anastomosis. We performed a double blind controlled trial to evaluate the safety and tolerance of recombinant human interleukin 10 (IL-10; Tenovil) in subjects operated on for Crohn's disease. We also assessed the effect of Tenovil in preventing endoscopic recurrence 12 weeks after surgery.
METHODS—Patients with Crohn's disease who underwent curative ileal or ileocolonic resection and primary anastomosis were randomised within two weeks after surgery to receive subcutaneous Tenovil 4 µg/kg once daily (QD) (n=22) or 8 µg/kg twice weekly (TIW) (n=21), or placebo (QD or TIW) (n=22). An ileocolonoscopy was performed after 12 weeks of treatment.
RESULTS—Compliance was excellent. The most frequently observed adverse events were mild and moderate in severity and equally distributed across treatment groups. Thirty seven patients in the pooled Tenovil group and 21 patients in the pooled placebo group were evaluable by endoscopy. At 12 weeks, 11 of 21 patients (52%) in the placebo group had recurrent lesions compared with 17 of 37 patients (46%) in the Tenovil group (ns). The incidence of severe endoscopic recurrence was similar in both groups (9%).
CONCLUSION—Tenovil treatment for 12 consecutive weeks in patients with Crohn's disease after intestinal resection was safe and well tolerated. No evidence of prevention of endoscopic recurrence of Crohn's disease by Tenovil was observed.


Keywords: Crohn's disease; interleukin 10; endoscopic recurrence PMID:11413109

  2. Interleukin 10 promoter region polymorphisms and susceptibility to advanced alcoholic liver disease

    PubMed Central

    Grove, J; Daly, A; Bassendine, M; Gilvarry, E; Day, C

    2000-01-01

    BACKGROUND—The factors determining why less than 10% of heavy drinkers develop advanced alcoholic liver disease (ALD) remain elusive, although genetic factors may be important. Interleukin 10 (IL-10) is an important cytokine with anti-inflammatory, anti-immune, and antifibrotic functions. Several polymorphisms have been identified in the IL-10 promoter and recent evidence suggests that some of these may have functional effects on IL-10 secretion.
AIMS—To test the hypothesis that IL-10 promoter region polymorphisms are associated with susceptibility to ALD.
METHODS—The allele frequencies for the two single base pair substitutions at positions −627 (C→A) and −1117 (A→G) in the IL-10 promoter were determined in 287 heavy drinkers with biopsy proved advanced ALD, 107 heavy drinkers with no evidence of liver disease or steatosis only on biopsy, and 227 local healthy volunteers.
RESULTS—At position −627, 50% of patients with advanced ALD had a least one A allele compared with 33% of controls (p<0.0001) and 34% of drinkers with no or mild disease (p=0.017). At position −1117, the slight excess of the A allele in drinkers with advanced disease was because of linkage disequilibrium between the A alleles at the two sites.
CONCLUSIONS—Among heavy drinkers, possession of the A allele at position −627 in the IL-10 promoter is associated with an increased risk of advanced liver disease. This is consistent with recent functional data that the −627*A allele is associated with low IL-10 expression which will favour inflammatory, immune mediated, and profibrotic mechanisms of alcohol related liver injury.


Keywords: ethyl alcohol; cirrhosis; interleukin 10; genetic polymorphism PMID:10716685

  3. Hypomethylation of proximal CpG motif of interleukin-10 promoter regulates its expression in human rheumatoid arthritis

    PubMed Central

    Fu, Li-hong; Ma, Chun-ling; Cong, Bin; Li, Shu-jin; Chen, Hai-ying; Zhang, Jing-ge

    2011-01-01

    Aim: The promoter of human interleukin-10 (IL10), a cytokine crucial for suppressing inflammation and regulating immune responses, contains an interspecies-conserved sequence with CpG motifs. The aim of this study was to investigate whether methylation of CpG motifs could regulate the expression of IL10 in rheumatoid arthritis (RA). Methods: Bioinformatic analysis was conducted to identify the interspecies-conserved sequence in human, macaque and mouse IL10 genes. Peripheral blood mononuclear cells (PBMCs) from 20 RA patients and 20 health controls were collected. The PBMCs from 6 patients were cultured in the presence or absence of 5-azacytidine (5 μmol/L). The mRNA and protein levels of IL10 were examined using RT-PCR and ELISA, respectively. The methylation of CpGs in the IL10 promoter was determined by pyrosequencing. Chromatin immunoprecipitation (ChIP) assays were performed to detect the cyclic AMP response element-binding protein (CREB)-DNA interactions. Results: One interspecies-conserved sequence was found within the IL10 promoter. The upstream CpGs at −408, −387, −385, and −355 bp were hypermethylated in PBMCs from both the RA patients and healthy controls. In contrast, the proximal CpG at −145 was hypomethylated to much more extent in the RA patients than in the healthy controls (P=0.016), which was correlated with higher IL10 mRNA and serum levels. In the 5-azacytidine-treated PBMCs, the CpG motifs were demethylated, and the expression levels of IL10 mRNA and protein was significantly increased. CHIP assays revealed increased phospho-CREB binding to the IL10 promoter. Conclusion: The methylation of the proximal CpGs in the IL10 promoter may regulate gene transcription in RA. PMID:21986577

  4. Transduction of interleukin-10 through renal artery attenuates vascular neointimal proliferation and infiltration of immune cells in rat renal allograft.

    PubMed

    Xie, Jingxin; Li, Xueyi; Meng, Dan; Liang, Qiujuan; Wang, Xinhong; Wang, Li; Wang, Rui; Xiang, Meng; Chen, Sifeng

    2016-08-01

    Renal transplantation is the treatment of choice for end-stage renal failure. Although acute rejection is not a major issue anymore, chronic rejection, especially vascular rejection, is still a major factor that might lead to allograft dysfunction on the long term. The role of the local immune-regulating cytokine interleukin-10 (IL-10) in chronic renal allograft is unclear. Many clinical observations showed that local IL-10 level was negatively related to kidney allograft function. It is unknown this negative relationship was the result of immunostimulatory property or insufficient immunosuppression property of local IL-10. We performed ex vivo transduction before transplantation through artery of the renal allograft using adeno-associated viral vectors carrying IL-10 gene. Twelve weeks after transplantation, we found intrarenal IL-10 gene transduction significantly inhibited arterial neointimal proliferation, the number of occluded intrarenal artery, interstitial fibrosis, peritubular capillary congestion and glomerular inflammation in renal allografts compared to control allografts receiving PBS or vectors carrying YFP. IL-10 transduction increased serum IL-10 level at 4 weeks but not at 8 and 12 weeks. Renal IL-10 level increased while serum creatinine decreased significantly in IL-10 group at 12 weeks compared to PBS or YFP controls. Immunohistochemical staining showed unchanged total T cells (CD3) and B cells (CD45R/B220), decreased cytotoxic T cells (CD8), macrophages (CD68) and increased CD4+ and FoxP3+ cells in IL-10 group. In summary, intrarenal IL-10 inhibited the allograft rejection while modulated immune response. PMID:27317647

  5. Gene therapy: Biological pacemaker created by gene transfer

    NASA Astrophysics Data System (ADS)

    Miake, Junichiro; Marbán, Eduardo; Nuss, H. Bradley

    2002-09-01

    The pacemaker cells of the heart initiate the heartbeat, sustain the circulation, and dictate the rate and rhythm of cardiac contraction. Circulatory collapse ensues when these specialized cells are damaged by disease, a situation that currently necessitates the implantation of an electronic pacemaker. Here we report the use of viral gene transfer to convert quiescent heart-muscle cells into pacemaker cells, and the successful generation of spontaneous, rhythmic electrical activity in the ventricle in vivo. Our results indicate that genetically engineered pacemakers could be developed as a possible alternative to implantable electronic devices.

  6. Association between Interleukin-10 gene polymorphisms and risk of early-onset preeclampsia

    PubMed Central

    Song, Limeng; Zhong, Mei

    2015-01-01

    We conducted a case-control study to investigate the role of IL-10 -1082A/G (rs1800896), -819T/C (rs1800871), and -592A/C (rs1800872) polymorphisms in the development of early-onset preeclampsia. Polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) assay was applied to assess the polymorphisms of IL-10 -1082A/G (rs1800896), -819T/C (rs1800871), and -592A/C (rs1800872). The genotype distributions of IL-10 -1082A/G (rs1800896), -819T/C (rs1800871), and -592A/C (rs1800872) confirmed with HWE in the controls, and the P value for HWE was 0.41, 0.38 and 0.26, respectively. The results of the multivariate logistic regression analysis revealed that the association of individuals expressing the CC genotype and AC+CC of IL-10 -592A/C (rs1800872) with a significantly increased risk of early-onset preeclampsia in co-dominant and dominant models, compared to the AA genotype; the OR (95% CI) for these individuals was determined to be 2.09 (1.12-3.90) and 1.66 (1.03-2.71), respectively. In the recessive model, we found that CC genotype of IL-10 -592A/C (rs1800872) was associated with the increased risk of early-onset preeclampsia when compared with AA+AC genotype (OR = 1.67; 95% CI = 1.01-2.92). In conclusion, our study has indicated that IL-10 -592A/C (rs1800872) polymorphism was associated with an increased risk of early-onset preeclampsia in a Chinese population. PMID:26617906

  7. Inhibition of experimental autoimmune uveoretinitis by systemic and subconjunctival adenovirus-mediated transfer of the viral IL-10 gene

    PubMed Central

    De Kozak, Y; Thillaye-Goldenberg, B; Naud, M -C; Viana Da Costa, A; Auriault, C; Verwaerde, C

    2002-01-01

    Pathological ocular manifestations result from a dysregulation in the balance between proinflammatory type 1 cytokines and regulatory type 2 cytokines. Interleukin-10 (IL-10) is an anti-inflammatory cytokine with potent immunosuppressive effects. We have examined the efficiency of viral IL-10 adenovirus (Ad-vIL-10)-mediated gene transfer on experimental autoimmune uveoretinitis (EAU) induced in mice and rats by purified retinal autoantigens, respectively, interphotoreceptor binding protein (IRBP) and S-antigen (S-Ag). B10-A mice that received a single unilateral injection of Ad-vIL-10 in the retro-orbital sinus venosus performed 1 day before immunization with IRBP in the footpads showed high levels of circulating vIL-10 in their sera and a significant reduction in pathological ocular manifestations. Lower levels of IFN-γ and IL-2 were found in cellular supernatants from IRBP-stimulated splenic cells in these treated mice. The local effect on ocular disease of vIL-10 was neutralized completely by injection of a monoclonal anti-vIL-10 antibody, demonstrating the specificity of the treatment. To determine whether the transfer of the vIL-10 gene within the periocular tissues of the eye could prevent acute EAU, a subconjunctival injection of Ad-vIL-10 was performed in Lewis rats simultaneously with S-antigen in the footpads. This injection determined in situ vIL-10 expression with very low circulating vIL-10 and led to a significant reduction of EAU without affecting the systemic immune response. The present results suggest that Ad-mediated gene transfer resulting in systemic and local expression of vIL-10 provide a promising approach for the treatment of uveitis. PMID:12390308

  8. Gene duplication and transfer events in plant mitochondria genome

    SciTech Connect

    Xiong Aisheng Peng Rihe; Zhuang Jing; Gao Feng; Zhu Bo; Fu Xiaoyan; Xue Yong; Jin Xiaofen; Tian Yongsheng; Zhao Wei; Yao Quanhong

    2008-11-07

    Gene or genome duplication events increase the amount of genetic material available to increase the genomic, and thereby phenotypic, complexity of organisms during evolution. Gene duplication and transfer events have been important to molecular evolution in all three domains of life, and may be the first step in the emergence of new gene functions. Gene transfer events have been proposed as another accelerator of evolution. The duplicated gene or genome, mainly nuclear, has been the subject of several recent reviews. In addition to the nuclear genome, organisms have organelle genomes, including mitochondrial genome. In this review, we briefly summarize gene duplication and transfer events in the plant mitochondrial genome.

  9. Optical gene transfer by femtosecond laser pulses

    NASA Astrophysics Data System (ADS)

    Konig, Karsten; Riemann, Iris; Tirlapur, Uday K.

    2003-07-01

    Targeted transfection of cells is an important technique for gene therapy and related biomedical applications. We delineate how high-intensity (1012 W/cm2) near-infrared (NIR) 80 MHz nanojoule femtosecond laser pulses can create highly localised membrane perforations within a minute focal volume, enabling non-invasive direct transfection of mammalian cells with DNA. We suspended Chinese hamster ovarian (CHO), rat kangaroo kidney epithelial (PtK2) and rat fibroblast cells in 0.5 ml culture medium in a sterile miniaturized cell chamber (JenLab GmbH, Jena, Germany) containing 0.2 μg plasmid DNA vector pEGFP-N1 (4.7 kb), which codes for green fluorescent protein (GFP). The NIR laser beam was introduced into a femtosecond laser scanning microscope (JenLab GmbH, Jena, Germany; focussed on the edge of the cell membrane of a target cell for 16 ms. The integration and expression efficiency of EGFP were assessed in situ by two-photon fluorescence-lifetime imaging using time-correlated single photon counting. The unique capability to transfer foreign DNA safely and efficiently into specific cell types (including stem cells), circumventing mechanical, electrical or chemical means, will have many applications, such as targeted gene therapy and DNA vaccination.

  10. Lentiviral vector gene transfer to porcine airways.

    PubMed

    Sinn, Patrick L; Cooney, Ashley L; Oakland, Mayumi; Dylla, Douglas E; Wallen, Tanner J; Pezzulo, Alejandro A; Chang, Eugene H; McCray, Paul B

    2012-01-01

    In this study, we investigated lentiviral vector development and transduction efficiencies in well-differentiated primary cultures of pig airway epithelia (PAE) and wild-type pigs in vivo. We noted gene transfer efficiencies similar to that observed for human airway epithelia (HAE). Interestingly, feline immunodeficiency virus (FIV)-based vectors transduced immortalized pig cells as well as pig primary cells more efficiently than HIV-1-based vectors. PAE express TRIM5α, a well-characterized species-specific lentiviral restriction factor. We contrasted the restrictive properties of porcine TRIM5α against FIV- and HIV-based vectors using gain and loss of function approaches. We observed no effect on HIV-1 or FIV conferred transgene expression in response to porcine TRIM5α overexpression or knockdown. To evaluate the ability of GP64-FIV to transduce porcine airways in vivo, we delivered vector expressing mCherry to the tracheal lobe of the lung and the ethmoid sinus of 4-week-old pigs. One week later, epithelial cells expressing mCherry were readily detected. Our findings indicate that pseudotyped FIV vectors confer similar tropisms in porcine epithelia as observed in human HAE and provide further support for the selection of GP64 as an appropriate envelope pseudotype for future preclinical gene therapy studies in the porcine model of cystic fibrosis (CF).Molecular Therapy - Nucleic Acids (2012) 1, e56; doi:10.1038/mtna.2012.47; published online 27 November 2012. PMID:23187455

  11. Preliminary Characterisation of Tumor Necrosis Factor Alpha and Interleukin-10 Responses to Chlamydia pecorum Infection in the Koala (Phascolarctos cinereus)

    PubMed Central

    Mathew, Marina; Beagley, Kenneth W.; Timms, Peter; Polkinghorne, Adam

    2013-01-01

    Debilitating infectious diseases caused by Chlamydia are major contributors to the decline of Australia's iconic native marsupial species, the koala (Phascolarctos cinereus). An understanding of koala chlamydial disease pathogenesis and the development of effective strategies to control infections continue to be hindered by an almost complete lack of species-specific immunological reagents. The cell-mediated immune response has been shown to play an influential role in the response to chlamydial infection in other hosts. The objective of this study, hence, was to provide preliminary data on the role of two key cytokines, pro-inflammatory tumour necrosis factor alpha (TNFα) and anti-inflammatory interleukin 10 (IL10), in the koala Chlamydia pecorum response. Utilising sequence homology between the cytokine sequences obtained from several recently sequenced marsupial genomes, this report describes the first mRNA sequences of any koala cytokine and the development of koala specific TNFα and IL10 real-time PCR assays to measure the expression of these genes from koala samples. In preliminary studies comparing wild koalas with overt chlamydial disease, previous evidence of C. pecorum infection or no signs of C. pecorum infection, we revealed strong but variable expression of TNFα and IL10 in wild koalas with current signs of chlamydiosis. The description of these assays and the preliminary data on the cell-mediated immune response of koalas to chlamydial infection paves the way for future studies characterising the koala immune response to a range of its pathogens while providing reagents to assist with measuring the efficacy of ongoing attempts to develop a koala chlamydial vaccine. PMID:23527290

  12. Plant transformation via pollen tube-mediated gene transfer

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Genetic transformation using foreign genes and the subsequent development of transgenic plants has been employed to develop enhanced elite germplasm. Although some skepticism exits regarding pollen tube-mediated gene transfer (PTT), reports demonstrating improved transformation efficiency with PTT ...

  13. A model of the ternary complex of interleukin-10 with its soluble receptors

    PubMed Central

    Pletnev, Sergei; Magracheva, Eugenia; Wlodawer, Alexander; Zdanov, Alexander

    2005-01-01

    Background Interleukin-10 (IL-10) is a cytokine whose main biological function is to suppress the immune response by induction of a signal(s) leading to inhibition of synthesis of a number of cytokines and their cellular receptors. Signal transduction is initiated upon formation of a ternary complex of IL-10 with two of its receptor chains, IL-10R1 and IL-10R2, expressed on the cell membrane. The affinity of IL-10R1 toward IL-10 is very high, which allowed determination of the crystal structure of IL-10 complexed with the extracellular/soluble domain of IL-10R1, while the affinity of IL-10R2 toward either IL-10 or IL-10/sIL-10R1 complex is quite low. This so far has prevented any attempts to obtain structural information about the ternary complex of IL-10 with its receptor chains. Results Structures of the second soluble receptor chain of interleukin-10 (sIL-10R2) and the ternary complex of IL-10/sIL-10R1/sIL-10R2 have been generated by homology modeling, which allowed us to identify residues involved in ligand-receptor and receptor-receptor interactions. Conclusion The previously experimentally determined structure of the intermediate/binary complex IL-10/sIL-10R1 is the same in the ternary complex. There are two binding sites for the second receptor chain on the surface of the IL-10/sIL-10R1 complex, involving both IL-10 and sIL-10R1. Most of the interactions are hydrophilic in nature, although each interface includes two internal hydrophobic clusters. The distance between C-termini of the receptor chains is 25 Å, which is common for known structures of ternary complexes of other cytokines. The structure is likely to represent the biologically active signaling complex of IL-10 with its receptor on the surface of the cell membrane. PMID:15985167

  14. Gene Transfer & Hybridization Studies in Hyperthermophilic Species

    SciTech Connect

    Nelson, Karen E.

    2005-10-14

    A. ABSTRACT The importance of lateral gene transfer (LGT) in the evolution of microbial species has become increasingly evident with each completed microbial genome sequence. Most significantly, the genome of Thermotoga maritima MSB8, a hyperthermophilic bacterium isolated by Karl Stetter and workers from Vulcano Italy in 1986, and sequenced at The Institute for Genomic Research (TIGR) in Rockville Maryland in 1999, revealed extensive LGT between % . this bacterium and members of the archaeal domain (in particular Archaeoglobus fulgidus, and Pyracoccus frcriosus species). Based on whole genome comparisons, it was estimated that 24% of the genetic information in this organism was acquired by genetic exchange with archaeal species, Independent analyses including periodicity analysis of the T. maritimu genomic DNA sequence, phylogenetic reconstruction based on genes that appear archaeal-like, and codon and amino acid usage, have provided additional evidence for LGT between T. maritima and the archaea. More recently, DiRuggiero and workers have identified a very recent LGT event between two genera of hyperthermophilic archaea, where a nearly identical DNA fragment of 16 kb in length flanked by insertion sequence (IS) elements, exists. Undoubtedly, additional examples of LGT will be identified as more microbial genomes are completed. For the present moment however, the genome sequence of T. maritima and other hyperthermophiles including P. furiosus, Pyrococcus horikoshii, Pyrococcus abyssi, A. fulgidus, and Aquifex aeolicus, have significantly increased out awareness of evolution being a web of life rather than a tree of life, as suggested by single gene phylogenies. In this proposal, we will aim to determine the extent of LGT across the hyperthemophiles, employing iY maritima as the model organism. A variety of biochemical techniques and phylogenetic reconstructions will allow for a detailed and thorough characterization of the extent of LGT in this species. The

  15. Interleukin-10 paradox: A potent immunoregulatory cytokine that has been difficult to harness for immunotherapy.

    PubMed

    Saxena, Ankit; Khosraviani, Sam; Noel, Sanjeev; Mohan, Divya; Donner, Thomas; Hamad, Abdel Rahim A

    2015-07-01

    Interleukin-10 (IL-10) is arguably the most potent anti-inflammatory cytokine. It is produced by almost all the innate and adaptive immune cells. These cells also serve as its targets, indicating that IL-10 secretion and action is highly regulated and perhaps compartmentalized. Consistent with this notion, various efforts directed at systemic administration of IL-10 to modulate autoimmune diseases (type 1 diabetes, multiple sclerosis, rheumatoid arthritis, psoriasis) have produced conflicting and largely inconsequential effects. On the other hand, IL-10 can promote humoral immune responses, enhancing class II expression on B cells and inducing immunoglobulin (Ig) production. Consequently, the high IL-10 level in systemic lupus erythematosus (SLE) patients is considered pathogenic and its blockade ameliorates the disease. In this perspective, we review preclinical findings and results of recent clinical studies using exogenous IL-10 to treat the aforementioned autoimmune diseases. In addition, given the limited success of IL-10 supplementation, we suggest that future studies should be expanded beyond modulating the delivery modes to include developing new strategies to protect and replenish the endogenous sources of IL-10. As an example, we provide evidence that aberrant Fas-mediated deletion of IL-10-producing B cells subverts the immunoregulatory role of IL-10 in autoimmune diabetes and that modulation of the Fas pathway preserves the IL-10-producing B cells and completely protects NOD mice from developing the disease. PMID:25481648

  16. Why interleukin-10 supplementation does not work in Crohn’s disease patients

    PubMed Central

    Marlow, Gareth J; van Gent, Dominique; Ferguson, Lynnette R

    2013-01-01

    Inflammatory bowel diseases (IBD) such as Crohn’s disease (CD) or ulcerative colitis are chronic intestinal disorders, which are on the increase in “Westernised” countries. IBD can be caused by both genetic and environmental factors. Interleukin-10 (IL-10) is an immunoregulatory cytokine that has been identified as being involved in several diseases including IBD. Studies have shown that polymorphisms in the promoter region reduce serum levels of IL-10 and this reduction has been associated with some forms of IBD. Mouse models have shown promising results with IL-10 supplementation, as such IL-10 supplementation has been touted as being a possible alternative treatment for CD in humans. Clinical trials have shown that recombinant human IL-10 is safe and well tolerated up to a dose of 8 μg/kg. However, to date, the results of the clinical trials have been disappointing. Although CD activity was reduced as measured by the CD activity index, IL-10 supplementation did not result in significantly reduced remission rates or clinical improvements when compared to placebo. This review discusses why IL-10 supplementation is not effective in CD patients currently and what can be addressed to potentially make IL-10 supplementation a more viable treatment option in the future. Based on the current research we conclude that IL-10 supplementation is not a one size fits all treatment and if the correct population of patients is chosen then IL-10 supplementation could be of benefit. PMID:23840137

  17. Epstein-Barr virus transformation induces B lymphocytes to produce human interleukin 10.

    PubMed

    Burdin, N; Péronne, C; Banchereau, J; Rousset, F

    1993-02-01

    Interleukin 10 (IL-10) is a pleiotropic factor that enhances proliferation of activated human B lymphocytes and induces them to secrete high amounts of immunoglobulins. Here we show that several human B cell lines were able to constitutively secrete human (h)IL-10. Whereas none of the pre-B nor the plasmocytic cell lines tested produced hIL-10, 25 of the 36 tested mature B cell lines (lymphoblastoid and Burkitt lymphoma cell lines) secreted hIL-10. Moreover, 24 of these 25 hIL-10-producing B cell lines contained the Epstein-Barr virus (EBV) genome, suggesting a relationship between hIL-10 production by human B cell lines and EBV expression. Accordingly, whereas polyclonal activation via triggering of surface immunoglobulins or CD40 antigen induced highly purified normal human B lymphocytes to produce only low (0.3-0.4 ng/ml) but significant amounts of hIL-10, EBV infection induced them to secrete high amounts of hIL-10 (4-9 ng/ml). Furthermore, addition of exogenous hIL-10, simultaneously to EBV infection, potentiated cell proliferation, whereas a blocking anti-IL-10 antiserum inhibited it. Thus, hIL-10 produced by infected human B lymphocytes appears to be involved in the mechanisms of EBV-induced B cell proliferation. PMID:8381152

  18. Interleukin-10 (IL-10) in Experimental Visceral Leishmaniasis and IL-10 Receptor Blockade as Immunotherapy

    PubMed Central

    Murray, Henry W.; Lu, Christina M.; Mauze, Smita; Freeman, Sherry; Moreira, Andre L.; Kaplan, Gilla; Coffman, Robert L.

    2002-01-01

    Interleukin-10 (IL-10) is thought to promote intracellular infection, including human visceral leishmaniasis, by disabling Th1 cell-type responses and/or deactivating parasitized tissue macrophages. To develop a rationale for IL-10 inhibition as treatment in visceral infection, Th1 cytokine-driven responses were characterized in Leishmania donovani-infected BALB/c mice in which IL-10 was absent or overexpressed or its receptor (IL-10R) was blockaded. IL-10 knockout and normal mice treated prophylactically with anti-IL-10R demonstrated accelerated granuloma assembly and rapid parasite killing without untoward tissue inflammation; IL-12 and gamma interferon mRNA expression, inducible nitric oxide synthase reactivity, and responsiveness to antimony chemotherapy were also enhanced in knockout mice. In IL-10 transgenic mice, parasite replication was unrestrained, and except for antimony responsiveness, measured Th1 cell-dependent events were all initially impaired. Despite subsequent granuloma assembly, high-level infection persisted, and antimony-treated transgenic mice also relapsed. In normal mice with established infection, anti-IL-10R treatment was remarkably active, inducing near-cure by itself and synergism with antimony. IL-10's deactivating effects regulate outcome in experimental visceral leishmaniasis, and IL-10R blockade represents a potential immuno- and/or immunochemotherapeutic approach in this infection. PMID:12379707

  19. Interleukin-10 inhibits lipopolysaccharide-induced upregulation of tissue factor in canine peripheral blood monocytes.

    PubMed

    Ogasawara, Seigo; Stokol, Tracy

    2012-08-15

    The potentially fatal hemostatic disorder of disseminated intravascular coagulation (DIC) is initiated in bacterial sepsis by lipopolysaccharide (LPS)-induced tissue factor (TF) expression on monocytes. Interleukin-10 (IL-10) is a potent inhibitory cytokine that downregulates monocyte inflammatory and procoagulant responses. We hypothesized that canine recombinant IL-10 (rIL-10) would inhibit LPS-induced TF upregulation on canine monocytes in a dose-dependent manner. Canine peripheral blood mononuclear cells (PBMC), obtained by double-density gradient centrifugation, and monocytes, purified from PBMC by immunomagnetic bead separation with an anti-canine CD14 antibody (Ab), were stimulated in suspension with LPS (0.1-1000 ng/mL) for various times. Recombinant IL-10 (10-5000 pg/mL) was added with LPS or up to 2h later. Tissue factor procoagulant activity was measured by cleavage of a chromogenic substrate by activated Factor X generated by the TF-factor VII complex. We found that rIL-10, when given concurrently or 1h after LPS, strongly inhibited LPS-induced TF procoagulant activity in canine PBMC and monocytes. This inhibition was dose-dependent and blocked by an anti-canine IL-10 Ab. Our results indicate that rIL-10 effectively inhibits LPS-induced TF upregulation in canine monocytes and could potentially be useful in limiting the development of DIC in dogs with endotoxemia. PMID:22609246

  20. Autoimmunity to heat shock protein 60 and antigen-specific production of interleukin-10.

    PubMed Central

    Yi, Y; Yang, X; Brunham, R C

    1997-01-01

    The immunopathologic sequelae of chlamydial infection are correlated with immune responses to the Chlamydia trachomatis heat shock protein 60 (hsp60). One pathogenic mechanism that may explain this association is the induction of autoimmune responses to self hsp60, since these two proteins share a high degree of amino acid sequence identity. To investigate the conditions under which autoimmune responses can be generated against self hsp60, groups of CBA mice were immunized with recombinant mouse hsp60, recombinant chlamydial hsp60, or both proteins. The data show that autoimmune responses characterized by strong T-cell proliferation and high titers of antibody to self hsp60 are induced only by concurrent immunization with mouse and chlamydial hsp60. Immunization with mouse hsp60 alone induced lymphocytes that secreted high levels of interleukin-10 (IL-10) but did not proliferate in response to in vitro stimulation with mouse hsp60; coimmunization with mouse and chlamydial hsp60s induced lymphocytes that proliferated strongly in response to mouse hsp60, secreted 6-fold less IL-10, and exhibited a 12-fold increase in the ratio of gamma interferon/IL-10 production. Switches in cytokine production patterns may mediate the pathogenesis of hsp60-associated diseases such as C. trachomatis immunopathology. PMID:9125545

  1. Hyporesponsiveness to the anti-inflammatory action of interleukin-10 in type 2 diabetes

    PubMed Central

    Barry, Julianne C.; Shakibakho, Soroush; Durrer, Cody; Simtchouk, Svetlana; Jawanda, Kamaldeep K.; Cheung, Sylvia T.; Mui, Alice L.; Little, Jonathan P.

    2016-01-01

    Chronic low-grade inflammation contributes to the pathology and complications of type 2 diabetes (T2D). Interleukin-10 (IL10), an anti-inflammatory cytokine, is suggested to play a protective role in T2D. However, the impact of T2D on IL10 function has not been previously assessed. We examined the ability of IL10 to inhibit inflammation in human T2D immune cells and explored underlying mechanisms using macrophage models. IL10 was less effective at inhibiting tumour necrosis factor (TNF)-α secretion in T2D whole blood cultures, which was not explained by altered IL10 receptor surface expression. These findings were observed in macrophages exposed to high glucose, which demonstrated similar IL10 resistance or hyporesponsiveness. These findings were also not explained by changes in IL10 receptor protein or other downstream signaling proteins. High glucose was also shown to impair the ability of IL10 to activate STAT3, a downstream signaling protein of IL10. Treatment with the SHIP1 agonist, AQX-MN100, reversed IL10 hyporesponsiveness in macrophages cultured in high glucose and showed equal effectiveness at different glucose conditions. This data supports the idea that IL10 hyporesponsiveness may contribute to chronic inflammation in T2D. These novel findings suggest that strategies aimed to overcome IL10 hyporesponsiveness may hold therapeutic potential for reducing inflammation in T2D. PMID:26883847

  2. Chemoenzymatic Synthesis of Nonasulfated Tetrahyaluronan with a Paramagnetic Tag for Studying Its Complex with Interleukin-10.

    PubMed

    Köhling, Sebastian; Künze, Georg; Lemmnitzer, Katharina; Bermudez, Marcel; Wolber, Gerhard; Schiller, Jürgen; Huster, Daniel; Rademann, Jörg

    2016-04-11

    Implants and artificial biomaterials containing sulfated hyaluronans have been shown to improve the healing of injured skin and bones. It is hypothesized that these effects are mediated by the binding of sulfated glycosaminoglycans (GAGs) to growth factors and cytokines, resulting in the sequestering of proteins to the wound healing site and in modulated protein activity. Given that no direct synthetic access to sulfated oligohyaluronans has been available, little is known about their protein binding and the structure of the resulting protein complexes. Here, the chemoenzymatic preparation of oligohyaluronans on the gram scale is described. Oligohyaluronans are converted into anomeric azides at the reducing end, enabling the attachment of analytical labels through an anomeric ligation reaction. A nonasulfated tetrahyaluronan-ethylenediaminetetraacetic acid derivative has been produced and used as a paramagnetic tag for the elucidation of the complex of this ligand with interleukin-10 using paramagnetic relaxation enhancement NMR analysis. The metal ion position is resolved with 1.0 Å, enabling a refined structural model of the complex. PMID:26918733

  3. Methods for Gene Transfer to the Central Nervous System

    PubMed Central

    Kantor, Boris; Bailey, Rachel M.; Wimberly, Keon; Kalburgi, Sahana N.; Gray, Steven J.

    2015-01-01

    Gene transfer is an increasingly utilized approach for research and clinical applications involving the central nervous system (CNS). Vectors for gene transfer can be as simple as an unmodified plasmid, but more commonly involve complex modifications to viruses to make them suitable gene delivery vehicles. This chapter will explain how tools for CNS gene transfer have been derived from naturally occurring viruses. The current capabilities of plasmid, retroviral, adeno-associated virus, adenovirus, and herpes simplex virus vectors for CNS gene delivery will be described. These include both focal and global CNS gene transfer strategies, with short- or long-term gene expression. As is described in this chapter, an important aspect of any vector is the cis-acting regulatory elements incorporated into the vector genome that control when, where, and how the transgene is expressed. PMID:25311922

  4. Gene transfer system for Rhodopseudomonas viridis.

    PubMed Central

    Lang, F S; Oesterhelt, D

    1989-01-01

    A gene transfer system for Rhodopseudomonas viridis was established which uses conjugation with Escherichia coli S17-I as the donor and mobilizable plasmids as vectors. Initially, plasmids of the incompatibility group P1 (pRK290 and pRK404) were used. The more effective shuttle vectors between E. coli and R. viridis, pKV1 and pKVS1, were derived from plasmid pBR322 and showed the highest conjugation frequency (10(-2] thus far demonstrated in purple bacteria. It was also demonstrated that Rhizobium meliloti can be used as a donor for conjugation with R. viridis. From a genomic cosmid library of R. viridis constructed in the vector pHC79, clones that coded for subunits H (puh operon), L, M and cytochrome c (puf operon) of the photosynthetic reaction center were isolated and characterized. For linkage of the two operons on the genome, cosmids that overlapped with the operon-carrying clones were identified. The relative positions of the two operons could not be determined, but the operons must be more than 100 kilobase pairs apart. Thus, the genomic organization of the reaction center in R. viridis is different from that of Rhodobacter capsulatus, for which a distance of about 39 kilobase pairs was determined. From a spontaneous mutant of R. viridis that is resistant to the herbicide terbutryn, the puf operon was cloned in pKVS1 and transferred by conjugation into R. viridis wild-type cells. The resulting exconjugants were resistant to the herbicide, which demonstrated that the puf operon on pKVS1 constructions was functionally expressed in R. viridis. Images PMID:2666398

  5. Problems associated with gene transfer and opportunities for microgravity environments

    NASA Astrophysics Data System (ADS)

    Tennessen, Daniel J.

    1997-01-01

    The method of crop improvement by gene transfer is becoming increasingly routine with transgenic foods and ornamental crops now being marketed to consumers. However, biological processes of plants, and the physical barriers of current protocols continue to limit the application of gene transfer in many commercial crops. The goal of this paper is to outline the current limitations of gene transfer and to hypothesize possible opportunities for use of microgravity to overcome such limitations. The limitations detailed in this paper include host-range specificity of Agrobacterium mediated transformation, probability of gene insertion, position effects of the inserted genes, gene copy number, stability of foreign gene expression in host plants, and regeneration of recalcitrant plant species. Microgravity offers an opportunity for gene transfer where cell growth kinetics, DNA synthesis, and genetic recombination rates can be altered. Such biological conditions may enhance the ability for recombination of reporter genes and other genes of interest to agriculture. Proposed studies would be useful for understanding instability of foreign gene expression and may lead to stable transformed plants. Other aspects of gene transfer in microgravity are discussed.

  6. Alterations of Colonic Contractility in an Interleukin-10 Knockout Mouse Model of Inflammatory Bowel Disease

    PubMed Central

    Park, Jae Hyung; Kwon, Joong Goo; Kim, Sun Joo; Song, Dae Kyu; Lee, Seok Guen; Kim, Eun Su; Cho, Kwang Bum; Jang, Byung Ik; Kim, Dae Hwan; Sin, Jeong-Im; Kim, Tae Wan; Song, In Hwan; Park, Kyung Sik

    2015-01-01

    Background/Aims Inflammatory bowel disease is commonly accompanied by colonic dysmotility and causes changes in intestinal smooth muscle contractility. In this study, colonic smooth muscle contractility in a chronic inflammatory condition was investigated using smooth muscle tissues prepared from interleukin-10 knockout (IL-10−/−) mice. Methods Prepared smooth muscle sections were placed in an organ bath system. Cholinergic and nitrergic neuronal responses were observed using carbachol and electrical field stimulation with L-NG-nitroarginine methyl ester (L-NAME). The expression of interstitial cells of Cajal (ICC) networks, muscarinic receptors, neuronal nitric oxide synthase (nNOS) and inducible nitric oxide synthase (iNOS) was observed via immunofluorescent staining. Results The spontaneous contractility and expression of ICC networks in the proximal and distal colon was significantly decreased in IL-10−/− mice compared to IL-10+/+ mice. The contractility in response to carbachol was significantly decreased in the proximal colon of IL-10−/− mice compared to IL-10+/+ mice, but no significant difference was found in the distal colon. In addition, the expression of muscarinic receptor type 2 was reduced in the proximal colon of IL-10−/− mice. The nictric oxide-mediated relaxation after electrical field stimulation was significantly decreased in the proximal and distal colon of IL-10−/− mice. In inflamed colon, the expression of nNOS decreased, whereas the expression of iNOS increased. Conclusions These results suggest that damage to the ICC network and NOS system in the proximal and distal colon, as well as damage to the smooth muscle cholinergic receptor in the proximal colon may play an important role in the dysmotility of the inflamed colon. PMID:25537671

  7. 3D domain swapping causes extensive multimerisation of human interleukin-10 when expressed in planta.

    PubMed

    Westerhof, Lotte B; Wilbers, Ruud H P; Roosien, Jan; van de Velde, Jan; Goverse, Aska; Bakker, Jaap; Schots, Arjen

    2012-01-01

    Heterologous expression platforms of biopharmaceutical proteins have been significantly improved over the last decade. Further improvement can be established by examining the intrinsic properties of proteins. Interleukin-10 (IL-10) is an anti-inflammatory cytokine with a short half-life that plays an important role in re-establishing immune homeostasis. This homodimeric protein of 36 kDa has significant therapeutic potential to treat inflammatory and autoimmune diseases. In this study we show that the major production bottleneck of human IL-10 is not protein instability as previously suggested, but extensive multimerisation due to its intrinsic 3D domain swapping characteristic. Extensive multimerisation of human IL-10 could be visualised as granules in planta. On the other hand, mouse IL-10 hardly multimerised, which could be largely attributed to its glycosylation. By introducing a short glycine-serine-linker between the fourth and fifth alpha helix of human IL-10 a stable monomeric form of IL-10 (hIL-10(mono)) was created that no longer multimerised and increased yield up to 20-fold. However, hIL-10(mono) no longer had the ability to reduce pro-inflammatory cytokine secretion from lipopolysaccharide-stimulated macrophages. Forcing dimerisation restored biological activity. This was achieved by fusing human IL-10(mono) to the C-terminal end of constant domains 2 and 3 of human immunoglobulin A (Fcα), a natural dimer. Stable dimeric forms of IL-10, like Fcα-IL-10, may not only be a better format for improved production, but also a more suitable format for medical applications. PMID:23049703

  8. Significance of decreased serum interleukin-10 levels in the progression of cerebral infarction.

    PubMed

    Diao, Zeng-Yan; Wang, Cui-Lan; Qi, Hong-Shun; Jia, Guo-Yong; Yan, Chuan-Zhu

    2016-05-01

    Anti-inflammatory cytokine and its serological detection may have an important role in the process of cardiovascular and cerebrovascular diseases. We investigated whether serum interleukin-10 (IL-10) is associated with cerebral infarction or not in the general population. Identified comprehensive searching was performed covering PubMed, EMBASE, Web of Science, Cochrane Library, CISCOM, CINAHL, Google Scholar, China BioMedicine, and China National Knowledge Infrastructure databases. Two reviewers extracted data and assessed studies independently. Information was extracted separately and classed into Asians and Caucasians. Summary standardized mean differences (SMDs) with 95 % confidence intervals (CI) were used with the utilization of Z test. Nine studies ranged from 2003 to 2014 were collected for meta-analysis. Results identified a negative association between serum IL-10 levels and cerebral infarction (SMD = 1.80, 95 % CI 0.79-2.81, P < 0.001). Country-subgroup analysis showed that low IL-10 level may be the main risk factor for cerebral infarction in India (SMD = 1.44, 95 % CI 1.13-1.75, P < 0.001) and Croatia (SMD = 2.96, 95 % CI 2.48-3.44, P < 0.001). In the ethnicity-stratified subgroup analysis, serum IL-10 levels were negatively correlated with cerebral infarction in Asians (SMD = 2.52, 95 % CI 0.47-4.57, P = 0.016), while not in Caucasians (P > 0.05). The lower serum IL-10 concentration was significantly associated with an increased likelihood of cerebral infarction in this meta-analysis. More prospective studies should be conducted to provide stronger evidence justifying the use of IL-10 as new biomarker to identify a predisposition toward cerebral infarction. PMID:25847570

  9. Regulator Versus Effector Paradigm: Interleukin-10 as Indicator of the Switching Response.

    PubMed

    Mingomataj, Ervin Ç; Bakiri, Alketa H

    2016-02-01

    The interleukin-10 (IL-10) is generally considered as the most important cytokine with anti-inflammatory properties and one of the key cytokines preventing inflammation-mediated tissue damage. In this respect, IL-10 producing cells play a crucial role in the outcome of infections, allergy, autoimmune reactions, tumor development, and transplant tolerance. Based on recent findings with regard to the mentioned clinical conditions, this review attempts to shed some light on the IL-10 functions, considering this cytokine as inherent inducer of the switching immunity. While acute infections and vaccinations are associated by IL-10 enhanced during few weeks, chronic parasitoses, tumor diseases, allergen-specific immunotherapy, transplants, and use of immune-suppressor drugs show an increased IL-10 level along months or years. With regard to autoimmune pathologies, the IL-10 increase is prevalently observed during early stages, whereas the successive stages are characterized by reaching of immune equilibrium independently to disease's activity. Together, these findings indicate that IL-10 is mainly produced during transient immune conditions and the persistent IL-10-related effect is the indication/prediction (and maybe effectuation) of the switching immunity. Actual knowledge emphasizes that any manipulation of the IL-10 response for treatment purposes should be considered very cautiously due to its potential hazards to the immune system. Probably, the IL-10 as potential switcher of immunity response should be used in association with co-stimulatory immune effectors that are necessary to determine the appropriate deviation during treatment of respective pathologies. Hopefully, further findings would open new avenues to study the biology of this "master switch" cytokine and its therapeutic potential. PMID:26450621

  10. Role of interleukin-10 (IL-10) in regulation of GABAergic transmission and acute response to ethanol.

    PubMed

    Suryanarayanan, A; Carter, J M; Landin, J D; Morrow, A L; Werner, D F; Spigelman, I

    2016-08-01

    Mounting evidence indicates that ethanol (EtOH) exposure activates neuroimmune signaling. Alterations in pro-inflammatory cytokines after acute and chronic EtOH exposure have been heavily investigated. In contrast, little is known about the regulation of neurotransmission and/or modulation by anti-inflammatory cytokines in the brain after an acute EtOH exposure. Recent evidence suggests that interleukin-10 (IL-10), an anti-inflammatory cytokine, is upregulated during withdrawal from chronic EtOH exposure. In the present study, we show that IL-10 is increased early (1 h) after a single intoxicating dose of EtOH (5 g/kg, intragastric) in Sprague Dawley rats. We also show that IL-10 rapidly regulates GABAergic transmission in dentate gyrus neurons. In brain slice recordings, IL-10 application dose-dependently decreases miniature inhibitory postsynaptic current (mIPSC) area and frequency, and decreases the magnitude of the picrotoxin sensitive tonic current (Itonic), indicating both pre- and postsynaptic mechanisms. A PI3K inhibitor LY294002 (but not the negative control LY303511) ablated the inhibitory effects of IL-10 on mIPSC area and Itonic, but not on mIPSC frequency, indicating the involvement of PI3K in postsynaptic effects of IL-10 on GABAergic transmission. Lastly, we also identify a novel neurobehavioral regulation of EtOH sensitivity by IL-10, whereby IL-10 attenuates acute EtOH-induced hypnosis. These results suggest that EtOH causes an early release of IL-10 in the brain, which may contribute to neuronal hyperexcitability as well as disturbed sleep seen after binge exposure to EtOH. These results also identify IL-10 signaling as a potential therapeutic target in alcohol-use disorders and other CNS disorders where GABAergic transmission is altered. PMID:27016017

  11. Targeted Interleukin-10 Nanotherapeutics Developed with a Microfluidic Chip Enhance Resolution of Inflammation in Advanced Atherosclerosis.

    PubMed

    Kamaly, Nazila; Fredman, Gabrielle; Fojas, Jhalique Jane R; Subramanian, Manikandan; Choi, Won Ii; Zepeda, Katherine; Vilos, Cristian; Yu, Mikyung; Gadde, Suresh; Wu, Jun; Milton, Jaclyn; Carvalho Leitao, Renata; Rosa Fernandes, Livia; Hasan, Moaraj; Gao, Huayi; Nguyen, Vance; Harris, Jordan; Tabas, Ira; Farokhzad, Omid C

    2016-05-24

    Inflammation is an essential protective biological response involving a coordinated cascade of signals between cytokines and immune signaling molecules that facilitate return to tissue homeostasis after acute injury or infection. However, inflammation is not effectively resolved in chronic inflammatory diseases such as atherosclerosis and can lead to tissue damage and exacerbation of the underlying condition. Therapeutics that dampen inflammation and enhance resolution are currently of considerable interest, in particular those that temper inflammation with minimal host collateral damage. Here we present the development and efficacy investigations of controlled-release polymeric nanoparticles incorporating the anti-inflammatory cytokine interleukin 10 (IL-10) for targeted delivery to atherosclerotic plaques. Nanoparticles were nanoengineered via self-assembly of biodegradable polyester polymers by nanoprecipitation using a rapid micromixer chip capable of producing nanoparticles with retained IL-10 bioactivity post-exposure to organic solvent. A systematic combinatorial approach was taken to screen nanoparticles, resulting in an optimal bioactive formulation from in vitro and ex vivo studies. The most potent nanoparticle termed Col-IV IL-10 NP22 significantly tempered acute inflammation in a self-limited peritonitis model and was shown to be more potent than native IL-10. Furthermore, the Col-IV IL-10 nanoparticles prevented vulnerable plaque formation by increasing fibrous cap thickness and decreasing necrotic cores in advanced lesions of high fat-fed LDLr(-/-) mice. These results demonstrate the efficacy and pro-resolving potential of this engineered nanoparticle for controlled delivery of the potent IL-10 cytokine for the treatment of atherosclerosis. PMID:27100066

  12. High-level production of human interleukin-10 fusions in tobacco cell suspension cultures.

    PubMed

    Kaldis, Angelo; Ahmad, Adil; Reid, Alexandra; McGarvey, Brian; Brandle, Jim; Ma, Shengwu; Jevnikar, Anthony; Kohalmi, Susanne E; Menassa, Rima

    2013-06-01

    The production of pharmaceutical proteins in plants has made much progress in recent years with the development of transient expression systems, transplastomic technology and humanizing glycosylation patterns in plants. However, the first therapeutic proteins approved for administration to humans and animals were made in plant cell suspensions for reasons of containment, rapid scale-up and lack of toxic contaminants. In this study, we have investigated the production of human interleukin-10 (IL-10) in tobacco BY-2 cell suspension and evaluated the effect of an elastin-like polypeptide tag (ELP) and a green fluorescent protein (GFP) tag on IL-10 accumulation. We report the highest accumulation levels of hIL-10 obtained with any stable plant expression system using the ELP fusion strategy. Although IL-10-ELP has cytokine activity, its activity is reduced compared to unfused IL-10, likely caused by interference of ELP with folding of IL-10. Green fluorescent protein has no effect on IL-10 accumulation, but examining the trafficking of IL-10-GFP over the cell culture cycle revealed fluorescence in the vacuole during the stationary phase of the culture growth cycle. Analysis of isolated vacuoles indicated that GFP alone is found in vacuoles, while the full-size fusion remains in the whole-cell extract. This indicates that GFP is cleaved off prior to its trafficking to the vacuole. On the other hand, IL-10-GFP-ELP remains mostly in the ER and accumulates to high levels. Protein bodies were observed at the end of the culture cycle and are thought to arise as a consequence of high levels of accumulation in the ER. PMID:23297698

  13. Interferon-alpha stimulates production of interleukin-10 in activated CD4+ T cells and monocytes.

    PubMed

    Aman, M J; Tretter, T; Eisenbeis, I; Bug, G; Decker, T; Aulitzky, W E; Tilg, H; Huber, C; Peschel, C

    1996-06-01

    In the present study, we investigated the effect of interferon-alpha (IFN-alpha) on the expression of interleukin-10 (IL-10) mRNA and protein synthesis in human monocytes and CD4+ T cells. In mononuclear cells, IFN-alpha induced expression of IL-10 mRNA and further enhanced lipopolysaccharide (LPS)-stimulated IL-10 expression. In purified monocytes, a strong expression of IL-10 mRNA induced by LPS was not further enhanced by IFN-alpha. In highly purified CD4+ T cells, IFN-alpha upregulated IL-10 mRNA upon activation with phytohemagglutinin and phorbol myristate acetate. In purified monocytes, an effect of IFN-alpha on IL-10 protein synthesis was dependent on costimulation with LPS. Maximal stimulation of IL-10 protein by IFN-alpha was seen after prolonged incubation periods of 48 to 96 hours, whereas IFN-gamma reduced IL-10 production in the early incubation period. Similar effects of IFN-alpha were observed in CD4+ T cells activated with CD3 and CD28 monoclonal antibodies. Addition of IFN-alpha caused an increase of IL-10 in culture supernatants of activated T-helper cells of more than 100% after 96 hours of incubation. In contrast, other cytokines, including IFN-gamma and IL-4, had no influence on IL-10 secretion stimulated by CD3 and CD28 in CD4+ T cells. In serum samples of IFN-alpha-treated individuals, we failed to detect an influence of cytokine treatment on IL-10 serum levels, confirming the requirement of additional activating signals for IFN-alpha-mediated effects on IL-10 synthesis. In conclusion, IFN-alpha enhances the late induction of IL-10, which physiologically occurs upon stimulation of monocytes and T cells. Biologically, this effect might enhance the negative-feedback mechanism ascribed to IL-10, which limits inflammatory reactions. PMID:8639843

  14. Anti-inflammatory cytokines in asthma and allergy: interleukin-10, interleukin-12, interferon-gamma.

    PubMed Central

    Chung, F

    2001-01-01

    Interleukin-10 (IL-10) is a cytokine derived from CD4+ T-helper type 2 (T(H2)) cells identified as a suppressor of cytokines from T-helper type 1(T(H1)) cells. Interleukin-12 (IL-12) is produced by B cells, macrophages and dendritic cells, and primarily regulates T(H1) cell differentiation, while suppressing the expansion of T(H2) cell clones. Interferon-gamma (IFN-gamma) is a product of T(H1) cells and exerts inhibitory effects on T(H2) cell differentiation. These cytokines have been implicated in the pathogenesis of asthma and allergies. In this context, IL-12 and IFN-gamma production in asthma have been found to be decreased, and this may reduce their capacity to inhibit IgE synthesis and allergic inflammation. IL-10 is a potent inhibitor of monocyte/macrophage function, suppressing the production of many pro-inflammatory cytokines. A relative underproduction of IL-10 from alveolar macrophages of atopic asthmatics has been reported. Therapeutic modulation of T(H1)/T(H2) imbalance in asthma and allergy by mycobacterial vaccine, specific immunotherapy and cytoline-guanosine dinucleotide motif may lead to increases in IL-12 and IFN-gamma production. Stimulation of IL-10 production by antigen-specific T-cells during immunotherapy may lead to anergy through inhibition of CD28-costimulatory molecule signalling by IL-10s anti-inflammatory effect on basophils, mast cells and eosinophils. PMID:11405550

  15. Exercise ameliorates high fat diet induced cardiac dysfunction by increasing interleukin 10

    PubMed Central

    Kesherwani, Varun; Chavali, Vishalakshi; Hackfort, Bryan T.; Tyagi, Suresh C.; Mishra, Paras K.

    2015-01-01

    Increasing evidence suggests that a sedentary lifestyle and a high fat diet (HFD) leads to cardiomyopathy. Moderate exercise ameliorates cardiac dysfunction, however underlying molecular mechanisms are poorly understood. Increased inflammation due to induction of pro-inflammatory cytokine such as tumor necrosis factor-alpha (TNF-α) and attenuation of anti-inflammatory cytokine such as interleukin 10 (IL-10) contributes to cardiac dysfunction in obese and diabetics. We hypothesized that exercise training ameliorates HFD- induced cardiac dysfunction by mitigating obesity and inflammation through upregulation of IL-10 and downregulation of TNF-α. To test this hypothesis, 8 week old, female C57BL/6J mice were fed with HFD and exercised (swimming 1 h/day for 5 days/week for 8 weeks). The four treatment groups: normal diet (ND), HFD, HFD + exercise (HFD + Ex) and ND + Ex were analyzed for mean body weight, blood glucose level, TNF-α, IL-10, cardiac fibrosis by Masson Trichrome, and cardiac dysfunction by echocardiography. Mean body weights were increased in HFD but comparatively less in HFD + Ex. The level of TNF-α was elevated and IL-10 was downregulated in HFD but ameliorated in HFD + Ex. Cardiac fibrosis increased in HFD and was attenuated by exercise in the HFD + Ex group. The percentage ejection fraction and fractional shortening were decreased in HFD but comparatively increased in HFD + Ex. There was no difference between ND and ND + Ex for the above parameters except an increase in IL-10 level following exercise. Based on these results, we conclude that exercise mitigates HFD- induced cardiomyopathy by decreasing obesity, inducing IL-10, and reducing TNF-α in mice. PMID:25954207

  16. Serum Levels of Interleukin-6 and Interleukin-10 as Biomarkers for Hepatocellular Carcinoma in Egyptian Patients

    PubMed Central

    Othman, Mohamed S.; Aref, Ahmed M.; Mohamed, Amal A.; Ibrahim, Wesam A.

    2013-01-01

    Interleukin-10 (IL-10) and interleukin-6 (IL-6) have been reported to be related to hepatocellular carcinoma (HCC) prognosis. This study aimed to investigate the clinical usefulness of serum levels of IL-6 and IL-10 as biomarkers for HCC among high-risk patients. Materials and Methods. 80 individuals were enrolled in this study; they were categorized into 4 groups: group 1 healthy individuals (NC) (n = 20), group 2 chronic hepatitis C virus (HCV) patients (n = 20), group 3 cirrhotic patients (LC) (n = 20), and HCC group (n = 20). Using ELISA technique serum levels of IL-6, IL-10, and alpha fetoprotein (AFP) were evaluated in all groups. Results. The mean serum levels of IL-6 were significantly higher in HCC than in LC, HCV, and NC groups (13.99 ± 1.80, 7.49 ± 0.43, 5.78 ± 0.74, and 2.57 ± 0.31), respectively (P < 0.05); also the serum levels of IL-10 were significantly higher in HCC compared with LC, HCV, and NC groups (13.69 ± 1.89, 7.37 ± 0.53, 5.18 ± 0.6, and 3.31 ± 0.42) (P < 0.05). We also found that the tumor size is correlated strongly with IL-6 and IL-10 levels (r = 0.925, P < 0.001; r = 0.821, P < 0.001), respectively. Conclusion. The combination of those markers may help to identify a group of HCC patients with low AFP.

  17. A recently transferred cluster of bacterial genes in Trichomonas vaginalis - lateral gene transfer and the fate of acquired genes

    PubMed Central

    2014-01-01

    Background Lateral Gene Transfer (LGT) has recently gained recognition as an important contributor to some eukaryote proteomes, but the mechanisms of acquisition and fixation in eukaryotic genomes are still uncertain. A previously defined norm for LGTs in microbial eukaryotes states that the majority are genes involved in metabolism, the LGTs are typically localized one by one, surrounded by vertically inherited genes on the chromosome, and phylogenetics shows that a broad collection of bacterial lineages have contributed to the transferome. Results A unique 34 kbp long fragment with 27 clustered genes (TvLF) of prokaryote origin was identified in the sequenced genome of the protozoan parasite Trichomonas vaginalis. Using a PCR based approach we confirmed the presence of the orthologous fragment in four additional T. vaginalis strains. Detailed sequence analyses unambiguously suggest that TvLF is the result of one single, recent LGT event. The proposed donor is a close relative to the firmicute bacterium Peptoniphilus harei. High nucleotide sequence similarity between T. vaginalis strains, as well as to P. harei, and the absence of homologs in other Trichomonas species, suggests that the transfer event took place after the radiation of the genus Trichomonas. Some genes have undergone pseudogenization and degradation, indicating that they may not be retained in the future. Functional annotations reveal that genes involved in informational processes are particularly prone to degradation. Conclusions We conclude that, although the majority of eukaryote LGTs are single gene occurrences, they may be acquired in clusters of several genes that are subsequently cleansed of evolutionarily less advantageous genes. PMID:24898731

  18. Cotransfer of linked eukaryotic genes and efficient transfer of hypoxanthine phosphoribosyltransferase by DNA-mediated gene transfer.

    PubMed Central

    Peterson, J L; McBride, O W

    1980-01-01

    The efficiency of DNA-mediated transfer of the gene (hprt) for hypoxanthine phosphoribosyltransferase (HPRT; IMP: pyrophosphate phosphoribosyltransferase, EC 2.4.2.8) is dependent upon the recipient cell used. hprt has been transferred into mouse TG8 or Chinese hamster CHTG49 cells at a high frequency, similar to the frequency of the gene (tk) for thymidine kinase (TK; ATP:thymidine 5'-phosphotransferase, EC 2.7.1.21) transfer into mouse LMTK- cells (i.e., 10(-6)). In contrast, the frequency of transfer of hprt into mouse A9 cells was about two orders of magnitude less. The identification of efficient recipient cells for hprt transfer permits the use of DNA-mediated transfer as a bioassay for the gene. Cotransfer of the linked tk gene and the gene (galk) for galactokinase (ATP: D-galactose 1-phosphotransferase, EC 2.7.1.6) to LMTK- cells has been detected once among 87 tk transferrents. This suggests that the distance between the tk and galk genes in the Chinese hamster genome may be smaller than was previously thought. Significant differences between chromosome-mediated and DNA-mediated gene transfer were observed with respect to both the size of the transferred functional genetic fragment and the recipient cell specificity. Images PMID:6929511

  19. Gene Transfer Strategies to Promote Chondrogenesis and Cartilage Regeneration.

    PubMed

    Im, Gun-Il

    2016-04-01

    Gene transfer has been used experimentally to promote chondrogenesis and cartilage regeneration. While it is controversial to apply gene therapy for nonlethal conditions such as cartilage defect, there is a possibility that the transfer of therapeutic transgenes may dramatically increase the effectiveness of cell therapy and reduce the quantity of cells that are needed to regenerate cartilage. Single or combination of growth factors and transcription factors has been transferred to mesenchymal stem cells or articular chondrocytes using both nonviral and viral approaches. The current challenge for the clinical applications of genetically modified cells is ensuring the safety of gene therapy while guaranteeing effectiveness. Viral gene delivery methods have been mainstays currently with enhanced safety features being recently refined. On the other hand, efficiency has been greatly improved in nonviral delivery. This review summarizes the history and recent update on the gene transfer to enhance chondrogenesis from stem cells or articular chondrocytes. PMID:26414246

  20. Mast cell-derived interleukin 10 limits skin pathology in contact dermatitis and chronic irradiation with ultraviolet B.

    PubMed

    Grimbaldeston, Michele A; Nakae, Susumu; Kalesnikoff, Janet; Tsai, Mindy; Galli, Stephen J

    2007-10-01

    Allergic contact dermatitis, such as in response to poison ivy or poison oak, and chronic low-dose ultraviolet B irradiation can damage the skin. Mast cells produce proinflammatory mediators that are thought to exacerbate these prevalent acquired immune or innate responses. Here we found that, unexpectedly, mast cells substantially limited the pathology associated with these responses, including infiltrates of leukocytes, epidermal hyperplasia and epidermal necrosis. Production of interleukin 10 by mast cells contributed to the anti-inflammatory or immunosuppressive effects of mast cells in these conditions. Our findings identify a previously unrecognized function for mast cells and mast cell-derived interleukin 10 in limiting leukocyte infiltration, inflammation and tissue damage associated with immunological or innate responses that can injure the skin. PMID:17767162

  1. Direct transfer of IL-12 gene into growing Renca tumors.

    PubMed

    Budryk, M; Wilczyńska, U; Szary, J; Szala, S

    2000-01-01

    We investigated the feasibility of transferring naked plasmid DNA containing a therapeutic gene (IL-12) into mice harboring growing Renca tumors. We found that naked DNA transferred into growing Renca and B16(F10) tumors gives higher expression level of reporter gene than complexes of DNA with DDAB/DOPE or DC-Chol/DOPE. Transfer of naked DNA carrying the IL-12 gene into growing Renca tumors causes a distinct therapeutic effect that depends on the time span between inoculation of mice with cancer cells and the beginning of the therapy. Therapy started on day 3 resulted in total cure (100%) of mice. PMID:11051203

  2. LATERAL GENE TRANSFER AND THE HISTORY OF BACTERIAL GENOMES

    SciTech Connect

    Howard Ochman

    2006-02-22

    The aims of this research were to elucidate the role and extent of lateral transfer in the differentiation of bacterial strains and species, and to assess the impact of gene transfer on the evolution of bacterial genomes. The ultimate goal of the project is to examine the dynamics of a core set of protein-coding genes (i.e., those that are distributed universally among Bacteria) by developing conserved primers that would allow their amplification and sequencing in any bacterial taxa. In addition, we adopted a bioinformatic approach to elucidate the extent of lateral gene transfer in sequenced genome.

  3. Intracellular gene transfer: Reduced hydrophobicity facilitates gene transfer for subunit 2 of cytochrome c oxidase

    PubMed Central

    Daley, Daniel O.; Clifton, Rachel; Whelan, James

    2002-01-01

    Subunit 2 of cytochrome c oxidase (Cox2) in legumes offers a rare opportunity to investigate factors necessary for successful gene transfer of a hydrophobic protein that is usually mitochondrial-encoded. We found that changes in local hydrophobicity were necessary to allow import of this nuclear-encoded protein into mitochondria. All legume species containing both a mitochondrial and nuclear encoded Cox2 displayed a similar pattern, with a large decrease in hydrophobicity evident in the first transmembrane region of the nuclear encoded protein compared with the organelle-encoded protein. Mitochondrial-encoded Cox2 could not be imported into mitochondria under the direction of the mitochondrial targeting sequence that readily supports the import of nuclear encoded Cox2. Removal of the first transmembrane region promotes import ability of the mitochondrial-encoded Cox2. Changing just two amino acids in the first transmembrane region of mitochondrial-encoded Cox2 to the corresponding amino acids in the nuclear encoded Cox2 also promotes import ability, whereas changing the same two amino acids in the nuclear encoded Cox2 to what they are in the mitochondrial-encoded copy prevents import. Therefore, changes in amino acids in the mature protein were necessary and sufficient for gene transfer to allow import under the direction of an appropriate signal to achieve the functional topology of Cox2. PMID:12142462

  4. Fibrin-mediated lentivirus gene transfer: implications for lentivirus microarrays

    PubMed Central

    Raut, Shruti; Lei, Pedro; Padmashali, Roshan; Andreadis, Stelios T.

    2010-01-01

    We employed fibrin hydrogel as bioactive matrix for lentivirus mediated gene transfer. Fibrin-mediated gene transfer was highly efficient and exhibited strong dependence on fibrinogen concentration. Efficient gene transfer was achieved with fibrinogen concentration between 3.75 – 7.5 mg/mL. Lower fibrinogen concentrations resulted in diffusion of virus out of the gel while higher concentrations led to ineffective fibrin degradation by target cells. Addition of fibrinolytic inhibitors decreased gene transfer in a dose-dependent manner suggesting that fibrin degradation by target cells may be necessary for successful gene delivery. Under these conditions transduction may be limited only to cells interacting with the matrix thereby providing a method for spatially localized gene delivery. Indeed, when lentivirus-containing fibrin microgels were spotted in an array format gene transfer was confined to virus-containing fibrin spots with minimal cross-contamination between neighboring sites. Collectively, our data suggest that fibrin may provide an effective matrix for spatially-localized gene delivery with potential applications in high-throughput lentiviral microarrays and in regenerative medicine. PMID:20153386

  5. Global Analysis of Horizontal Gene Transfer in Fusarium verticillioides

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The co-occurrence of microbes within plants and other specialized niches may facilitate horizontal gene transfer (HGT) affecting host-pathogen interactions. We recently identified fungal-to-fungal HGTs involving metabolic gene clusters. For a global analysis of HGTs in the maize pathogen Fusarium ve...

  6. Evolution of and horizontal gene transfer in the Endornavirus genus.

    PubMed

    Song, Dami; Cho, Won Kyong; Park, Sang-Ho; Jo, Yeonhwa; Kim, Kook-Hyung

    2013-01-01

    The transfer of genetic information between unrelated species is referred to as horizontal gene transfer. Previous studies have demonstrated that both retroviral and non-retroviral sequences have been integrated into eukaryotic genomes. Recently, we identified many non-retroviral sequences in plant genomes. In this study, we investigated the evolutionary origin and gene transfer of domains present in endornaviruses which are double-stranded RNA viruses. Using the available sequences for endornaviruses, we found that Bell pepper endornavirus-like sequences homologous to the glycosyltransferase 28 domain are present in plants, fungi, and bacteria. The phylogenetic analysis revealed the glycosyltransferase 28 domain of Bell pepper endornavirus may have originated from bacteria. In addition, two domains of Oryza sativa endornavirus, a glycosyltransferase sugar-binding domain and a capsular polysaccharide synthesis protein, also exhibited high similarity to those of bacteria. We found evidence that at least four independent horizontal gene transfer events for the glycosyltransferase 28 domain have occurred among plants, fungi, and bacteria. The glycosyltransferase sugar-binding domains of two proteobacteria may have been horizontally transferred to the genome of Thalassiosira pseudonana. Our study is the first to show that three glycome-related viral genes in the genus Endornavirus have been acquired from marine bacteria by horizontal gene transfer. PMID:23667703

  7. Evolution of and Horizontal Gene Transfer in the Endornavirus Genus

    PubMed Central

    Park, Sang-Ho; Jo, Yeonhwa; Kim, Kook-Hyung

    2013-01-01

    The transfer of genetic information between unrelated species is referred to as horizontal gene transfer. Previous studies have demonstrated that both retroviral and non-retroviral sequences have been integrated into eukaryotic genomes. Recently, we identified many non-retroviral sequences in plant genomes. In this study, we investigated the evolutionary origin and gene transfer of domains present in endornaviruses which are double-stranded RNA viruses. Using the available sequences for endornaviruses, we found that Bell pepper endornavirus-like sequences homologous to the glycosyltransferase 28 domain are present in plants, fungi, and bacteria. The phylogenetic analysis revealed the glycosyltransferase 28 domain of Bell pepper endornavirus may have originated from bacteria. In addition, two domains of Oryza sativa endornavirus, a glycosyltransferase sugar-binding domain and a capsular polysaccharide synthesis protein, also exhibited high similarity to those of bacteria. We found evidence that at least four independent horizontal gene transfer events for the glycosyltransferase 28 domain have occurred among plants, fungi, and bacteria. The glycosyltransferase sugar-binding domains of two proteobacteria may have been horizontally transferred to the genome of Thalassiosira pseudonana. Our study is the first to show that three glycome-related viral genes in the genus Endornavirus have been acquired from marine bacteria by horizontal gene transfer. PMID:23667703

  8. Impact of iron overload on interleukin-10 levels, biochemical parameters and oxidative stress in patients with sickle cell anemia

    PubMed Central

    Barbosa, Maritza Cavalcante; dos Santos, Talyta Ellen Jesus; de Souza, Geane Félix; de Assis, Lívia Coêlho; Freitas, Max Victor Carioca; Gonçalves, Romélia Pinheiro

    2013-01-01

    Objective The aim of this study was to evaluate the impact of iron overload on the profile of interleukin-10 levels, biochemical parameters and oxidative stress in sickle cell anemia patients. Methods A cross-sectional study was performed of 30 patients with molecular diagnosis of sickle cell anemia. Patients were stratified into two groups, according to the presence of iron overload: Iron overload (n = 15) and Non-iron overload (n = 15). Biochemical analyses were performed utilizing the Wiener CM 200 automatic analyzer. The interleukin-10 level was measured by capture ELISA using the BD OptEIAT commercial kit. Oxidative stress parameters were determined by spectrophotometry. Statistical analysis was performed using GraphPad Prism software (version 5.0) and statistical significance was established for p-values < 0.05 in all analyses. Results Biochemical analysis revealed significant elevations in the levels of uric acid, triglycerides, very low-density lipoprotein (VLDL), alanine aminotransferase (ALT), lactate dehydrogenase (LDH), urea and creatinine in the Iron overload Group compared to the Non-iron overload Group and significant decreases in the high-density lipoprotein (HDL) and low-density lipoprotein (LDL). Ferritin levels correlated positively with uric acid concentrations (p-value < 0.05). The Iron overload Group showed lower interleukin-10 levels and catalase activity and higher nitrite and malondialdehyde levels compared with the Non-iron overload Group. Conclusion The results of this study are important to develop further consistent studies that evaluate the effect of iron overload on the inflammatory profile and oxidative stress of patients with sickle cell anemia. PMID:23580881

  9. Horizontal functional gene transfer from bacteria to fishes

    PubMed Central

    Sun, Bao-Fa; Li, Tong; Xiao, Jin-Hua; Jia, Ling-Yi; Liu, Li; Zhang, Peng; Murphy, Robert W.; He, Shun-Min; Huang, Da-Wei

    2015-01-01

    Invertebrates can acquire functional genes via horizontal gene transfer (HGT) from bacteria but fishes are not known to do so. We provide the first reliable evidence of one HGT event from marine bacteria to fishes. The HGT appears to have occurred after emergence of the teleosts. The transferred gene is expressed and regulated developmentally. Its successful integration and expression may change the genetic and metabolic repertoire of fishes. In addition, this gene contains conserved domains and similar tertiary structures in fishes and their putative donor bacteria. Thus, it may function similarly in both groups. Evolutionary analyses indicate that it evolved under purifying selection, further indicating its conserved function. We document the first likely case of HGT of functional gene from prokaryote to fishes. This discovery certifies that HGT can influence vertebrate evolution. PMID:26691285

  10. Interleukin-10 prevents epithelial cell apoptosis by regulating IFNγ and TNFα expression in rhesus macaque colon explants

    PubMed Central

    Pan, Diganta; Das, Arpita; Lala, Wendy; Kenway-Lynch, Carys S.; Liu, David X.; Veazey, Ronald S.; Pahar, Bapi

    2013-01-01

    Interleukin-10 (IL-10) is an important immunomodulatory cytokine that plays an obligate role in regulating inflammatory responses. Here we demonstrated the role of IL-10 in regulating crypts length and breadth as well as maintaining the survival of epithelial cells using rhesus colon explant cultures. Anti-IL-10 antibody treatment of colon explant cultures induced increased production of inflammatory cytokines/molecules like IFNγ, TNFα, CD107a and perforin as well as increased epithelial cell apoptosis compared to media controls tested. Our results suggest that IL-10 plays a crucial role in maintaining mucosal homeostasis by regulating mucosal IFNγ and TNFα cytokine production. PMID:23867612

  11. Interleukin-10 prevents epithelial cell apoptosis by regulating IFNγ and TNFα expression in rhesus macaque colon explants.

    PubMed

    Pan, Diganta; Das, Arpita; Lala, Wendy; Kenway-Lynch, Carys S; Liu, David X; Veazey, Ronald S; Pahar, Bapi

    2013-10-01

    Interleukin-10 (IL-10) is an important immunomodulatory cytokine that plays an obligate role in regulating inflammatory responses. Here we demonstrated the role of IL-10 in regulating crypts length and breadth as well as maintaining the survival of epithelial cells using rhesus colon explant cultures. Anti-IL-10 antibody treatment of colon explant cultures induced increased production of inflammatory cytokines/molecules like IFNγ, TNFα, CD107a and perforin as well as increased epithelial cell apoptosis compared to media controls tested. Our results suggest that IL-10 plays a crucial role in maintaining mucosal homeostasis by regulating mucosal IFNγ and TNFα cytokine production. PMID:23867612

  12. Advancements in gene transfer-based therapy for hemophilia A

    PubMed Central

    Doering, Christopher B; Spencer, H Trent

    2010-01-01

    Gene therapy has promised clinical benefit to those suffering with hemophilia A, but this benefit has not yet been realized. However, during the past two decades, basic and applied gene therapy research has progressed and the goal of gene therapy for hemophilia A is once again in our sights. The hemophilia A patient population suffers from a disease that requires invasive, lifelong management, is exorbitantly expensive to treat, has geographically limited treatment access and can become untreatable due to immune reactions to the treatment product. Subsequent to the cloning of the factor VIII gene and cDNA in the early 1980s, academic and commercial research laboratories began to pursue gene transfer-based therapies to supplement or supplant the available protein replacement therapy. However, to date, clinical trials for gene therapy of hemophilia A have been unsuccessful. Three trials have been conducted with each having tested a different gene-transfer strategy and each demonstrating that there is a considerable barrier to achieving sustained expression of therapeutic amounts of factor VIII. Recent progress has been made in gene-transfer technology and, relevant to hemophilia A, towards increasing the biosynthetic efficiency of factor VIII. These advances are now being combined to develop novel strategies to treat and possibly cure hemophilia A. PMID:20577574

  13. Analysis of Polymorphisms in Interleukin-10, Interleukin-6, and Interleukin-1 Receptor Antagonist in Mexican-Mestizo Women with Pre-eclampsia

    PubMed Central

    Valencia Villalvazo, Elith Yazmin; Canto-Cetina, Thelma; Romero Arauz, Juan Fernando; Coral-Vázquez, Ramón Mauricio; Canizales-Quinteros, Samuel; Coronel, Agustín; Carlos Falcón, Juan; Hernández Rivera, Jaime; Ibarra, Roberto; Polanco Reyes, Lucila

    2012-01-01

    Due to the fact that studies seeking associations of polymorphisms in regulatory regions of cytokine genes with pre-eclampsia (PE) have not always been consistent in different population analyses, the aim of this study was to investigate the possible association between rs1800896 of interleukin-10 (IL-10), rs1800795 of interleukin-6 (IL-6), and the variable number of tandem repeats (VNTR) in intron 2 of interleukin-1 receptor antagonist (IL-1Ra), as well as gene–gene interactions between these three polymorphisms with the presence of PE in Mexican-Mestizo women and one Amerindian population from México (Maya). A case–control study was performed where 411 pre-eclamptic cases and 613 controls were genotyped. For the rs1800896 of IL-10 and rs1800795 of IL-6, we used real-time polymerase chain reaction (PCR) allelic discrimination and for the VNTR of IL-1Ra, PCR. Allele frequency differences were assessed by Chi-squared test; logistic regression was used to test for associations; a gene–gene interaction was conducted. Genotypic and allelic distribution of the polymorphisms was similar in our population. The estimated of the gene–gene interaction between the polymorphisms did not differ significantly. However, we observed important differences in the distribution of the alleles and genotypes of the three polymorphisms analyzed between Mestiza-Mexicanas and Maya-Mestizo women. In conclusion, we did not find an association between polymorphisms in IL-10, IL-6, and IL-1Ra and PE in Mexican-Mestizo and Maya-Mestizo women. To our knowledge, this is the first time that these three polymorphisms were analyzed together with gene–gene interaction in women with PE. PMID:23013217

  14. Rational Structure-Based Rescaffolding Approach to De Novo Design of Interleukin 10 (IL-10) Receptor-1 Mimetics

    PubMed Central

    Philipp, Jenny; Künze, Georg; Wodtke, Robert; Löser, Reik; Fahmy, Karim; Pisabarro, M. Teresa

    2016-01-01

    Tackling protein interfaces with small molecules capable of modulating protein-protein interactions remains a challenge in structure-based ligand design. Particularly arduous are cases in which the epitopes involved in molecular recognition have a non-structured and discontinuous nature. Here, the basic strategy of translating continuous binding epitopes into mimetic scaffolds cannot be applied, and other innovative approaches are therefore required. We present a structure-based rational approach involving the use of a regular expression syntax inspired in the well established PROSITE to define minimal descriptors of geometric and functional constraints signifying relevant functionalities for recognition in protein interfaces of non-continuous and unstructured nature. These descriptors feed a search engine that explores the currently available three-dimensional chemical space of the Protein Data Bank (PDB) in order to identify in a straightforward manner regular architectures containing the desired functionalities, which could be used as templates to guide the rational design of small natural-like scaffolds mimicking the targeted recognition site. The application of this rescaffolding strategy to the discovery of natural scaffolds incorporating a selection of functionalities of interleukin-10 receptor-1 (IL-10R1), which are relevant for its interaction with interleukin-10 (IL-10) has resulted in the de novo design of a new class of potent IL-10 peptidomimetic ligands. PMID:27123592

  15. Rational Structure-Based Rescaffolding Approach to De Novo Design of Interleukin 10 (IL-10) Receptor-1 Mimetics.

    PubMed

    Ruiz-Gómez, Gloria; Hawkins, John C; Philipp, Jenny; Künze, Georg; Wodtke, Robert; Löser, Reik; Fahmy, Karim; Pisabarro, M Teresa

    2016-01-01

    Tackling protein interfaces with small molecules capable of modulating protein-protein interactions remains a challenge in structure-based ligand design. Particularly arduous are cases in which the epitopes involved in molecular recognition have a non-structured and discontinuous nature. Here, the basic strategy of translating continuous binding epitopes into mimetic scaffolds cannot be applied, and other innovative approaches are therefore required. We present a structure-based rational approach involving the use of a regular expression syntax inspired in the well established PROSITE to define minimal descriptors of geometric and functional constraints signifying relevant functionalities for recognition in protein interfaces of non-continuous and unstructured nature. These descriptors feed a search engine that explores the currently available three-dimensional chemical space of the Protein Data Bank (PDB) in order to identify in a straightforward manner regular architectures containing the desired functionalities, which could be used as templates to guide the rational design of small natural-like scaffolds mimicking the targeted recognition site. The application of this rescaffolding strategy to the discovery of natural scaffolds incorporating a selection of functionalities of interleukin-10 receptor-1 (IL-10R1), which are relevant for its interaction with interleukin-10 (IL-10) has resulted in the de novo design of a new class of potent IL-10 peptidomimetic ligands. PMID:27123592

  16. Horizontal gene transfer in the human gastrointestinal tract: potential spread of antibiotic resistance genes

    PubMed Central

    Huddleston, Jennifer R

    2014-01-01

    Bacterial infections are becoming increasingly difficult to treat due to widespread antibiotic resistance among pathogens. This review aims to give an overview of the major horizontal transfer mechanisms and their evolution and then demonstrate the human lower gastrointestinal tract as an environment in which horizontal gene transfer of resistance determinants occurs. Finally, implications for antibiotic usage and the development of resistant infections and persistence of antibiotic resistance genes in populations as a result of horizontal gene transfer in the large intestine will be discussed. PMID:25018641

  17. Experiments on gene transferring to primary hematopoietic cells by liposome.

    PubMed

    Hu, L; Zhang, B

    2000-01-01

    Liposomes have showed many advantages in mediating exogenous gene into many cell types in vitro and in vivo. But few data are available concerning gene transfer into hematopoietic cells. In this report, we described two-marker genes (Neo R and Lac Z) co-transferred into hematopoietic cells of human and mouse by using liposome in vitro. The efficiency of gene transfer was tested by X-gal staining and observation of colony formation. The X-gal blue staining rate of transduced cells was about (13.33 +/- 2.68)% in human and about (16.28 +/- 2.95)% in mouse without G418 selection. After G418 selection, the blue cell rate was (46.06 +/- 3.47)% in human and (43.45 +/- 4.1)% in mouse, which were markedly higher than those before selection, suggesting that high-efficiency gene transfer and expression could be attained in primary hematopoietic cells using this easy and harmless transduction protocol. At the same time, this protocol provided experimental data for clinicians to investigate the biology of marrow reconstitution and trace the origin of relapse after autologous bone marrow transplantation for the patients with leukemia. PMID:12840913

  18. Estimating the Frequency of Horizontal Gene Transfer Using Phylogenetic Models of Gene Gain and Loss.

    PubMed

    Zamani-Dahaj, Seyed Alireza; Okasha, Mohamed; Kosakowski, Jakub; Higgs, Paul G

    2016-07-01

    We analyze patterns of gene presence and absence in a maximum likelihood framework with rate parameters for gene gain and loss. Standard methods allow independent gains and losses in different parts of a tree. While losses of the same gene are likely to be frequent, multiple gains need to be considered carefully. A gene gain could occur by horizontal transfer or by origin of a gene within the lineage being studied. If a gene is gained more than once, then at least one of these gains must be a horizontal transfer. A key parameter is the ratio of gain to loss rates, a/v We consider the limiting case known as the infinitely many genes model, where a/v tends to zero and a gene cannot be gained more than once. The infinitely many genes model is used as a null model in comparison to models that allow multiple gains. Using genome data from cyanobacteria and archaea, it is found that the likelihood is significantly improved by allowing for multiple gains, but the average a/v is very small. The fraction of genes whose presence/absence pattern is best explained by multiple gains is only 15% in the cyanobacteria and 20% and 39% in two data sets of archaea. The distribution of rates of gene loss is very broad, which explains why many genes follow a treelike pattern of vertical inheritance, despite the presence of a significant minority of genes that undergo horizontal transfer. PMID:27189546

  19. Horizontal gene transfer in eukaryotes: The weak-link model

    PubMed Central

    Huang, Jinling

    2013-01-01

    The significance of horizontal gene transfer (HGT) in eukaryotic evolution remains controversial. Although many eukaryotic genes are of bacterial origin, they are often interpreted as being derived from mitochondria or plastids. Because of their fixed gene pool and gene loss, however, mitochondria and plastids alone cannot adequately explain the presence of all, or even the majority, of bacterial genes in eukaryotes. Available data indicate that no insurmountable barrier to HGT exists, even in complex multicellular eukaryotes. In addition, the discovery of both recent and ancient HGT events in all major eukaryotic groups suggests that HGT has been a regular occurrence throughout the history of eukaryotic evolution. A model of HGT is proposed that suggests both unicellular and early developmental stages as likely entry points for foreign genes into multicellular eukaryotes. PMID:24037739

  20. Viral mediated gene transfer to sprouting blood vessels during angiogenesis.

    PubMed

    Alian, Akram; Eldor, Amiram; Falk, Haya; Panet, Amos

    2002-08-01

    Several experimental systems have been applied to investigate the development of new blood vessels. Angiogenesis can be followed ex-vivo by culturing explants of rat aorta 'rings' in biomatrix gels. This angiogenesis system was modified for the study of viral vector mediated gene transfer, using adenovirus, vaccinia- and retroviral vectors. Two modifications were introduced to the model in order to facilitate efficient viral mediated gene transfer, (i) placing the aorta ring on top of a thin layer of collagen such that the angiogenic tissue will be accessible to the viral vector; and (ii) infection of the aorta rings prior to embedding them into the collagen matrix. While adenovirus and vaccinia vectors infected efficiently the aorta rings they induced cell death. Subsequent gene transfer experiments were, therefore, carried with retroviral vectors containing vascular endothelial growth factor (VEGF) and the beta-interferon (IFN) genes. Overexpression of VEGF enhanced significantly microvessel sprouting, while overexpression of IFN-beta induced an antiviral effect. The experimental system described in this study can facilitate the application of other viral vectors to the study of genes that may regulate the complex angiogenic process and thereby open new avenues for vascular gene therapy. PMID:12176137

  1. Gene Transfer in Mycobacterium tuberculosis: Shuttle Phasmids to Enlightenment

    PubMed Central

    JACOBS, WILLIAM R.

    2016-01-01

    Infectious diseases have plagued humankind throughout history and have posed serious public health problems. Yet vaccines have eradicated smallpox and antibiotics have drastically decreased the mortality rate of many infectious agents. These remarkable successes in the control of infections came from knowing the causative agents of the diseases, followed by serendipitous discoveries of attenuated viruses and antibiotics. The discovery of DNA as genetic material and the understanding of how this information translates into specific phenotypes have changed the paradigm for developing new vaccines, drugs, and diagnostic tests. Knowledge of the mechanisms of immunity and mechanisms of action of drugs has led to new vaccines and new antimicrobial agents. The key to the acquisition of the knowledge of these mechanisms has been identifying the elemental causes (i.e., genes and their products) that mediate immunity and drug resistance. The identification of these genes is made possible by being able to transfer the genes or mutated forms of the genes into causative agents or surrogate hosts. Such an approach was limited in Mycobacterium tuberculosis by the difficulty of transferring genes or alleles into M. tuberculosis or a suitable surrogate mycobacterial host. The construction of shuttle phasmids—chimeric molecules that replicate in Escherichia coli as plasmids and in mycobacteria as mycobacteriophages—was instrumental in developing gene transfer systems for M. tuberculosis. This review will discuss M. tuberculosis genetic systems and their impact on tuberculosis research. “I had to know my enemy in order to prevail against him.”Nelson Mandela PMID:26105819

  2. Genetic, epidemiological and biological analysis of interleukin-10 promoter single-nucleotide polymorphisms suggests a definitive role for -819C/T in leprosy susceptibility.

    PubMed

    Pereira, A C; Brito-de-Souza, V N; Cardoso, C C; Dias-Baptista, I M F; Parelli, F P C; Venturini, J; Villani-Moreno, F R; Pacheco, A G; Moraes, M O

    2009-03-01

    Leprosy is a complex infectious disease influenced by genetic and environmental factors. The genetic contributing factors are considered heterogeneous and several genes have been consistently associated with susceptibility like PARK2, tumor necrosis factor (TNF), lymphotoxin-alpha (LTA) and vitamin-D receptor (VDR). Here, we combined a case-control study (374 patients and 380 controls), with meta-analysis (5 studies; 2702 individuals) and biological study to test the epidemiological and physiological relevance of the interleukin-10 (IL-10) genetic markers in leprosy. We observed that the -819T allele is associated with leprosy susceptibility either in the case-control or in the meta-analysis studies. Haplotypes combining promoter single-nucleotide polymorphisms also implicated a haplotype carrying the -819T allele in leprosy susceptibility (odds ratio (OR)=1.40; P=0.01). Finally, we tested IL-10 production in peripheral blood mononuclear cells stimulated with Mycobacterium leprae antigens and found that -819T carriers produced lower levels of IL-10 when compared with non-carriers. Taken together, these data suggest that low levels of IL-10 during the disease outcome can drive patients to a chronic and unprotective response that culminates with leprosy. PMID:19110537

  3. Detecting Horizontal Gene Transfer between Closely Related Taxa.

    PubMed

    Adato, Orit; Ninyo, Noga; Gophna, Uri; Snir, Sagi

    2015-10-01

    Horizontal gene transfer (HGT), the transfer of genetic material between organisms, is crucial for genetic innovation and the evolution of genome architecture. Existing HGT detection algorithms rely on a strong phylogenetic signal distinguishing the transferred sequence from ancestral (vertically derived) genes in its recipient genome. Detecting HGT between closely related species or strains is challenging, as the phylogenetic signal is usually weak and the nucleotide composition is normally nearly identical. Nevertheless, there is a great importance in detecting HGT between congeneric species or strains, especially in clinical microbiology, where understanding the emergence of new virulent and drug-resistant strains is crucial, and often time-sensitive. We developed a novel, self-contained technique named Near HGT, based on the synteny index, to measure the divergence of a gene from its native genomic environment and used it to identify candidate HGT events between closely related strains. The method confirms candidate transferred genes based on the constant relative mutability (CRM). Using CRM, the algorithm assigns a confidence score based on "unusual" sequence divergence. A gene exhibiting exceptional deviations according to both synteny and mutability criteria, is considered a validated HGT product. We first employed the technique to a set of three E. coli strains and detected several highly probable horizontally acquired genes. We then compared the method to existing HGT detection tools using a larger strain data set. When combined with additional approaches our new algorithm provides richer picture and brings us closer to the goal of detecting all newly acquired genes in a particular strain. PMID:26439115

  4. Detecting Horizontal Gene Transfer between Closely Related Taxa

    PubMed Central

    Adato, Orit; Ninyo, Noga; Gophna, Uri; Snir, Sagi

    2015-01-01

    Horizontal gene transfer (HGT), the transfer of genetic material between organisms, is crucial for genetic innovation and the evolution of genome architecture. Existing HGT detection algorithms rely on a strong phylogenetic signal distinguishing the transferred sequence from ancestral (vertically derived) genes in its recipient genome. Detecting HGT between closely related species or strains is challenging, as the phylogenetic signal is usually weak and the nucleotide composition is normally nearly identical. Nevertheless, there is a great importance in detecting HGT between congeneric species or strains, especially in clinical microbiology, where understanding the emergence of new virulent and drug-resistant strains is crucial, and often time-sensitive. We developed a novel, self-contained technique named Near HGT, based on the synteny index, to measure the divergence of a gene from its native genomic environment and used it to identify candidate HGT events between closely related strains. The method confirms candidate transferred genes based on the constant relative mutability (CRM). Using CRM, the algorithm assigns a confidence score based on “unusual” sequence divergence. A gene exhibiting exceptional deviations according to both synteny and mutability criteria, is considered a validated HGT product. We first employed the technique to a set of three E. coli strains and detected several highly probable horizontally acquired genes. We then compared the method to existing HGT detection tools using a larger strain data set. When combined with additional approaches our new algorithm provides richer picture and brings us closer to the goal of detecting all newly acquired genes in a particular strain. PMID:26439115

  5. Specific Gene Repression by CRISPRi System Transferred through Bacterial Conjugation

    PubMed Central

    2014-01-01

    In microbial communities, bacterial populations are commonly controlled using indiscriminate, broad range antibiotics. There are few ways to target specific strains effectively without disrupting the entire microbiome and local environment. Here, we use conjugation, a natural DNA horizontal transfer process among bacterial species, to deliver an engineered CRISPR interference (CRISPRi) system for targeting specific genes in recipient Escherichia coli cells. We show that delivery of the CRISPRi system is successful and can specifically repress a reporter gene in recipient cells, thereby establishing a new tool for gene regulation across bacterial cells and potentially for bacterial population control. PMID:25409531

  6. Antibiotics and gene transfer in swine gut bacteria

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The mammalian gastrointestinal (GI) tract hosts a diverse collection bacteria, most of which are beneficial for host health. This bacterial community also supports a community of viruses that infect bacteria (called bacteriophages or phages). Phages transfer genes between bacteria, and phage-media...

  7. Transferred interbacterial antagonism genes augment eukaryotic innate immune function

    PubMed Central

    Chou, Seemay; Daugherty, Matthew D.; Peterson, S. Brook; Biboy, Jacob; Yang, Youyun; Jutras, Brandon L.; Fritz-Laylin, Lillian K.; Ferrin, Michael A.; Harding, Brittany N.; Jacobs-Wagner, Christine; Yang, X. Frank; Vollmer, Waldemar; Malik, Harmit S.

    2015-01-01

    Horizontal gene transfer (HGT) allows organisms to rapidly acquire adaptive traits1. Though documented instances of HGT from bacteria to eukaryotes remain rare, bacteria represent a rich source of new functions potentially available for co-option2. One benefit that genes of bacterial origin could provide to eukaryotes is the capacity to produce anti-bacterials, which have evolved in prokaryotes as the result of eons of interbacterial competition. The type VI secretion amidase effector (Tae) proteins are potent bacteriocidal enzymes that degrade the cell wall when delivered into competing bacterial cells by the type VI secretion system (T6SS)3. Here we show that tae genes have been transferred to eukaryotes on at least six occasions, and that the resulting domesticated amidase effector (dae) genes have been preserved for hundreds of millions of years via purifying selection. We show that the dae genes acquired eukaryotic secretion signals, are expressed within recipient organisms, and encode active antibacterial toxins that possess substrate specificity matching extant Tae proteins of the same lineage. Finally, we show that a dae gene in the deer tick Ixodes scapularis limits proliferation of Borrelia burgdorferi, the etiologic agent of Lyme disease. Our work demonstrates that a family of horizontally acquired toxins honed to mediate interbacterial antagonism confers previously undescribed antibacterial capacity to eukaryotes. We speculate that the selective pressure imposed by competition between bacteria has produced a reservoir of genes encoding diverse antimicrobial functions that are tailored for facile co-option by eukaryotic innate immune systems. PMID:25470067

  8. Detection of horizontal gene transfers from phylogenetic comparisons.

    PubMed

    Pylro, Victor Satler; Vespoli, Luciano de Souza; Duarte, Gabriela Frois; Yotoko, Karla Suemy Clemente

    2012-01-01

    Bacterial phylogenies have become one of the most important challenges for microbial ecology. This field started in the mid-1970s with the aim of using the sequence of the small subunit ribosomal RNA (16S) tool to infer bacterial phylogenies. Phylogenetic hypotheses based on other sequences usually give conflicting topologies that reveal different evolutionary histories, which in some cases may be the result of horizontal gene transfer events. Currently, one of the major goals of molecular biology is to understand the role that horizontal gene transfer plays in species adaptation and evolution. In this work, we compared the phylogenetic tree based on 16S with the tree based on dszC, a gene involved in the cleavage of carbon-sulfur bonds. Bacteria of several genera perform this survival task when living in environments lacking free mineral sulfur. The biochemical pathway of the desulphurization process was extensively studied due to its economic importance, since this step is expensive and indispensable in fuel production. Our results clearly show that horizontal gene transfer events could be detected using common phylogenetic methods with gene sequences obtained from public sequence databases. PMID:22675653

  9. Rescuing the Failing Heart by Targeted Gene Transfer

    PubMed Central

    Kawase, Yoshiaki; Ladage, Dennis; Hajjar, Roger J.

    2011-01-01

    Congestive heart failure is a major cause of morbidity and mortality in the US. While progress in conventional treatments is making steady and incremental gains to reduce heart failure mortality, there is a critical need to explore new therapeutic approaches. Gene therapy was initially applied in the clinical setting for inherited monogenic disorders. It is now apparent that gene therapy has broader potential that also includes acquired polygenic diseases, such as congestive heart failure. Recent advances in understanding of the molecular basis of myocardial dysfunction, together with the evolution of increasingly efficient gene transfer technology, has placed heart failure within reach of gene-based therapy. Furthermore, the recent successful and safe completion of a phase 2 trial targeting the sarcoplasmic reticulum calcium ATPase pump (SERCA2a) along with the start of more recent phase 1 trials usher a new era for gene therapy for the treatment of heart failure. PMID:21371634

  10. AAV-mediated gene transfer to the mouse CNS

    PubMed Central

    Stoica, Lorelei; Ahmed, Seemin S.

    2013-01-01

    Recombinant adeno associated virus (rAAV) vectors are great tools for gene transfer due to their ability to mediate long-term gene expression. Recombinant AAVs have been used at various ages of development with no apparent toxicity. There are multiple ways of delivering AAV vectors to the CNS, depending on the stage of development of the mouse. In neonates, intravascular injections into the facial vein are often used. In adults, direct injections into target regions of the brain are achieved with great spatiotemporal control through stereotaxic surgeries. Recently, discoveries of new AAV vectors with the ability to cross the blood brain barrier have made it possible to also target the adult CNS by intravascular injections. rAAVs have been successfully used as gene transfer vehicles in multiple animal models of CNS disorders, and several clinical trials are currently underway. PMID:23686825

  11. Endosymbiotic gene transfer from prokaryotic pangenomes: Inherited chimerism in eukaryotes

    PubMed Central

    Ku, Chuan; Nelson-Sathi, Shijulal; Roettger, Mayo; Garg, Sriram; Hazkani-Covo, Einat; Martin, William F.

    2015-01-01

    Endosymbiotic theory in eukaryotic-cell evolution rests upon a foundation of three cornerstone partners—the plastid (a cyanobacterium), the mitochondrion (a proteobacterium), and its host (an archaeon)—and carries a corollary that, over time, the majority of genes once present in the organelle genomes were relinquished to the chromosomes of the host (endosymbiotic gene transfer). However, notwithstanding eukaryote-specific gene inventions, single-gene phylogenies have never traced eukaryotic genes to three single prokaryotic sources, an issue that hinges crucially upon factors influencing phylogenetic inference. In the age of genomes, single-gene trees, once used to test the predictions of endosymbiotic theory, now spawn new theories that stand to eventually replace endosymbiotic theory with descriptive, gene tree-based variants featuring supernumerary symbionts: prokaryotic partners distinct from the cornerstone trio and whose existence is inferred solely from single-gene trees. We reason that the endosymbiotic ancestors of mitochondria and chloroplasts brought into the eukaryotic—and plant and algal—lineage a genome-sized sample of genes from the proteobacterial and cyanobacterial pangenomes of their respective day and that, even if molecular phylogeny were artifact-free, sampling prokaryotic pangenomes through endosymbiotic gene transfer would lead to inherited chimerism. Recombination in prokaryotes (transduction, conjugation, transformation) differs from recombination in eukaryotes (sex). Prokaryotic recombination leads to pangenomes, and eukaryotic recombination leads to vertical inheritance. Viewed from the perspective of endosymbiotic theory, the critical transition at the eukaryote origin that allowed escape from Muller’s ratchet—the origin of eukaryotic recombination, or sex—might have required surprisingly little evolutionary innovation. PMID:25733873

  12. Endosymbiotic gene transfer from prokaryotic pangenomes: Inherited chimerism in eukaryotes.

    PubMed

    Ku, Chuan; Nelson-Sathi, Shijulal; Roettger, Mayo; Garg, Sriram; Hazkani-Covo, Einat; Martin, William F

    2015-08-18

    Endosymbiotic theory in eukaryotic-cell evolution rests upon a foundation of three cornerstone partners--the plastid (a cyanobacterium), the mitochondrion (a proteobacterium), and its host (an archaeon)--and carries a corollary that, over time, the majority of genes once present in the organelle genomes were relinquished to the chromosomes of the host (endosymbiotic gene transfer). However, notwithstanding eukaryote-specific gene inventions, single-gene phylogenies have never traced eukaryotic genes to three single prokaryotic sources, an issue that hinges crucially upon factors influencing phylogenetic inference. In the age of genomes, single-gene trees, once used to test the predictions of endosymbiotic theory, now spawn new theories that stand to eventually replace endosymbiotic theory with descriptive, gene tree-based variants featuring supernumerary symbionts: prokaryotic partners distinct from the cornerstone trio and whose existence is inferred solely from single-gene trees. We reason that the endosymbiotic ancestors of mitochondria and chloroplasts brought into the eukaryotic--and plant and algal--lineage a genome-sized sample of genes from the proteobacterial and cyanobacterial pangenomes of their respective day and that, even if molecular phylogeny were artifact-free, sampling prokaryotic pangenomes through endosymbiotic gene transfer would lead to inherited chimerism. Recombination in prokaryotes (transduction, conjugation, transformation) differs from recombination in eukaryotes (sex). Prokaryotic recombination leads to pangenomes, and eukaryotic recombination leads to vertical inheritance. Viewed from the perspective of endosymbiotic theory, the critical transition at the eukaryote origin that allowed escape from Muller's ratchet--the origin of eukaryotic recombination, or sex--might have required surprisingly little evolutionary innovation. PMID:25733873

  13. Control of pro-inflammatory cytokine release from human monocytes with the use of an interleukin-10 monoclonal antibody.

    PubMed

    Patel, Hardik; Davidson, Dennis

    2014-01-01

    The monocytes (MONOs) can be considered as "double-edge swords"; they have both important pro-inflammatory and anti-inflammatory functions manifested in part by cytokine production and release. Although MONOs are circulating cells, they are the major precursors of a variety of tissue-specific immune cells such as the alveolar macrophage, dendritic cells, microglial cells, and Kupffer cells. Unlike the polymorphonuclear leukocyte, which produces no or very little interleukin-10 (IL-10), the monocyte can produce this potent anti-inflammatory cytokine to control inflammation. IL-10, on an equimolar basis, is a more potent inhibitor of pro-inflammatory cytokines produced by monocytes than many anti-inflammatory glucocorticoids which are used clinically. This chapter describes how to isolate monocytes from human blood and the use of IL-10 monoclonal antibody to determine the effect and timing of endogenous IL-10 release on the production and release of pro-inflammatory cytokines. PMID:24908297

  14. The cytomegalovirus homolog of interleukin-10 requires phosphatidylinositol 3-kinase activity for inhibition of cytokine synthesis in monocytes.

    PubMed

    Spencer, Juliet V

    2007-02-01

    Human cytomegalovirus (CMV) has evolved numerous strategies for evading host immune defenses, including piracy of cellular cytokines. A viral homolog of interleukin-10, designated cmvIL-10, binds to the cellular IL-10 receptor and effects potent immune suppression. The signaling pathways employed by cmvIL-10 were investigated, and the classic IL-10R/JAK1/Stat3 pathway was found to be activated in monocytes. However, inhibition of JAK1 had little effect on cmvIL-10-mediated suppression of tumor necrosis factor alpha (TNF-alpha) production. Inhibition of the phosphatidylinositol 3-kinase/Akt pathway had a more significant impact on TNF-alpha levels but did not completely relieve the immune suppression, demonstrating that cmvIL-10 stimulates multiple signaling pathways to modulate cell function. PMID:17121792

  15. Human gene transfer: Characterization of human tumor-infiltrating lymphocytes as vehicles for retroviral-mediated gene transfer in man

    SciTech Connect

    Kasid, A.; Morecki, S.; Aebersold, P.; Cornetta, K.; Culver, K.; Freeman, S.; Director, E.; Lotze, M.T.; Blaese, R.M.; Anderson, W.F.; Rosenberg, S.A. )

    1990-01-01

    Tumor-infiltrating lymphocytes (TILs) are cells generated from tumor suspensions cultured in interleukin 2 that can mediate cancer regression when adoptively transferred into mice or humans. Since TILs proliferate rapidly in vitro, recirculate, and preferentially localize at the tumor site in vivo, they provide an attractive model for delivery of exogenous genetic material into man. To determine whether efficient gene transfer into TILs is feasible. The authors transduced human TILs with the bacterial gene for neomycin-resistance (Neo{sup R}) using the retroviral vector N2. The transduced TIL populations were stable and polyclonal with respect to the intact Neo{sup R} gene integration and expressed high levels of neomycin phosphotransferase activity. The Neo{sup R} gene insertion did not alter the in vitro growth pattern and interleukin 2 dependence of the transduced TILs. Analyses of T-cell receptor gene rearrangement for {beta}- and {gamma}-chain genes revealed the oligoclonal nature of the TIL populations with no major change in the DNA rearrangement patterns or the levels of mRNA expression of the {beta} and {gamma} chains following transduction and selection of TILs in the neomycin analog G418. Human TILs expressed mRNA for tumor necrosis factors ({alpha} and {beta}) and interleukin 2 receptor P55. This pattern of cytokine-mRNA expression was not significantly altered following the transduction of TILs. The studies demonstrate the feasibility of TILs as suitable cellular vehicles for the introduction of therapeutic genes into patients receiving autologous TILs.

  16. Immunotherapy of Malignancy by in vivo Gene Transfer into Tumors

    NASA Astrophysics Data System (ADS)

    Plautz, Gregory E.; Yang, Zhi-Yong; Wu, Bei-Yue; Gao, Xiang; Huang, Leaf; Nabel, Gary J.

    1993-05-01

    The immune system confers protection against a variety of pathogens and contributes to the surveillance and destruction of neoplastic cells. Several cell types participate in the recognition and lysis of tumors, and appropriate immune stimulation provides therapeutic effects in malignancy. Foreign major histocompatibility complex (MHC) proteins also serve as a potent stimulus to the immune system. In this report, a foreign MHC gene was introduced directly into malignant tumors in vivo in an effort to stimulate tumor rejection. In contrast to previous attempts to induce tumor immunity by cell-mediated gene transfer, the recombinant gene was introduced directly into tumors in vivo. Expression of the murine class I H-2K^s gene within the CT26 mouse colon adenocarcinoma (H-2K^d) or the MCA 106 fibrosarcoma (H-2K^b) induced a cytotoxic T-cell response to H-2K^s and, more importantly, to other antigens present on unmodified tumor cells. This immune response attenuated tumor growth and caused complete tumor regression in many cases. Direct gene transfer in vivo can therefore induce cell-mediated immunity against specific gene products, which provides an immunotherapeutic effect for malignancy, and potentially can be applied to the treatment of cancer and infectious diseases in man.

  17. [Experimental study of vascular gene transfer using soluble stent].

    PubMed

    Huang, Z; Gou, J; Li, X

    1997-05-01

    We assessed the possibility of gene transfer into anastomotic arteries in vivo using soluble stent containing Adv5-CMV/LacZ. After being soaked in a high concentration solution of glucose containing Adv5-CMV (control group) or Adv5-CMV/LacZ (treatment group) for 30 minutes, we inserted soluble stents into the lumina of cut rat carotid arteries, and end-to-end anastomoses of cut rat carotid were performed with standard microvascular surgical technique. 16 rats were killed after two weeks, the segments of anastomotic carotid arteries were prepared for assessing beta-galactosidase activity and histochemical staining. In the control group, the anastomotic arteries did not have detectable level of beta-galactosidase expression. In the treatment group, the amount of beta-galactosidase expression was 9.80 x 10-3 u/g tissue. Microscopic examination of histochemically stained arteries demonstrated that gene transfered not only to endothelial cells but also to smooth muscle cells, and all anastomotic arteries were transfered in the treatment group, but none of arteries revealed blue in the control group. The results of this experimental study suggested that soluble stent be a new method of direct gene transfer into arteries in vivo. PMID:10374573

  18. Interleukin-10 overexpression does not synergize with the neuroprotective action of RGD-containing vectors after postnatal brain excitotoxicity but modulates the main inflammatory cell responses.

    PubMed

    Gonzalez, Pau; Peluffo, Hugo; Acarin, Laia; Villaverde, Antonio; Gonzalez, Berta; Castellano, Bernardo

    2012-01-01

    Antiinflammatory cytokines such as interleukin-10 (IL-10) have been used to modulate and terminate inflammation and provide neuroprotection. Recently, we reported that the modular recombinant transfection vector NLSCt is an efficient tool for transgene overexpression in vivo, which induces neuroprotection as a result of its RGD-mediated integrin-interacting capacity. We here sought to evaluate the putative synergic neuroprotective action exerted by IL-10 overexpression using NLSCt as a transfection vector after an excitotoxic injury to the postnatal rat brain. For this purpose, lesion volume, neurodegeneration, astroglial and microglial responses, neutrophil infiltration, and proinflammatory cytokine production were analyzed at several survival times after intracortical NMDA injection in postnatal day 9 rats, followed by injection of NLSCt combined with the IL-10 gene, a control transgene, or saline vehicle solution. Our results show no combined neuroprotective effect between RGD-interacting vectors and IL-10 gene therapy; instead, IL-10 overexpression using NLSCt as transfection vector increased lesion volume and neuronal degeneration at 12 hr and 3 days postlesion. In parallel, NLSCt/IL-10 treated animals displayed increased density of neutrophils and microglia/macrophages, and a reduced astroglial content of GFAP and vimentin. Moreover, NLSCt/IL-10 treated animals did not show any variation in interleukin-1β or tumor necrosis factor-α expression but a slight increase in interleukin-6 content at 7 days postlesion. In conclusion, overexpression of IL-10 by using NLSCt transfection vector did not synergistically neuroprotect the excitotoxically damaged postnatal rat brain but induced changes in the astroglial and microglial and inflammatory cell response. PMID:21922521

  19. Nano-Sized Sunflower Polycations As Effective Gene Transfer Vehicles.

    PubMed

    Cheng, Yilong; Wei, Hua; Tan, James-Kevin Y; Peeler, David J; Maris, Don O; Sellers, Drew L; Horner, Philip J; Pun, Suzie H

    2016-05-01

    The architecture of polycations plays an important role in both gene transfection efficiency and cytotoxicity. In this work, a new polymer, sunflower poly(2-dimethyl amino)ethyl methacrylate) (pDMAEMA), is prepared by atom transfer radical polymerization and employed as nucleic acid carriers compared to linear pDMAEMA homopolymer and comb pDMAEMA. The sunflower pDMAEMAs show higher IC50 , greater buffering capacity, and stronger binding capacity toward plasmid DNA than their linear and comb counterparts. In vitro transfection studies demonstrate that sunflower pDMAEMAs exhibit high transfection efficiency as well as relatively low cytotoxicity in complete growth medium. In vivo gene delivery by intraventricular injection to the brain shows that sunflower polymer delivers plasmid DNA more effectively than comb polymer. This study provides a new insight into the relationship between polymeric architecture and gene delivery capability, and as well as a useful means to design potent vectors for successful gene delivery. PMID:27061622

  20. Electroporation-Mediated Gene Transfer Directly to the Swine Heart

    PubMed Central

    Hargrave, Barbara; Downey, Harre; Strange, Robert; Murray, Len; Cinnamond, Cade; Lundberg, Cathryn; Israel, Annelise; Chen, Yeong-Jer; Marshall, William; Heller, Richard

    2012-01-01

    In vivo gene transfer to the ischemic heart via electroporation holds promise as a potential therapeutic approach for the treatment of heart disease. In the current study, we investigated the use of in vivo electroporation for gene transfer using 3 different penetrating electrodes and one non-penetrating electrode. The hearts of adult male swine were exposed through a sternotomy. Eight electric pulses synchronized to the rising phase of the R wave of the ECG were administered at varying pulse widths and field strengths following an injection of either a plasmid encoding luciferase or one encoding green fluorescent protein. Four sites on the anterior wall of the left ventricle were treated. Animals were euthanized 48 hours after injection and electroporation and gene expression was determined. Results were compared to sites in the heart that received plasmid injection but no electric pulses or were not treated. Gene expression was higher in all electroporated sites when compared to injection only sites demonstrating the robustness of this approach. Our results provide evidence that in vivo electroporation can be a safe and effective non-viral method for delivering genes to the heart, in vivo. PMID:22456328

  1. Characterization of an Ancient Lepidopteran Lateral Gene Transfer

    PubMed Central

    Wheeler, David; Redding, Amanda J.; Werren, John H.

    2013-01-01

    Bacteria to eukaryote lateral gene transfers (LGT) are an important potential source of material for the evolution of novel genetic traits. The explosion in the number of newly sequenced genomes provides opportunities to identify and characterize examples of these lateral gene transfer events, and to assess their role in the evolution of new genes. In this paper, we describe an ancient lepidopteran LGT of a glycosyl hydrolase family 31 gene (GH31) from an Enterococcus bacteria. PCR amplification between the LGT and a flanking insect gene confirmed that the GH31 was integrated into the Bombyx mori genome and was not a result of an assembly error. Database searches in combination with degenerate PCR on a panel of 7 lepidopteran families confirmed that the GH31 LGT event occurred deep within the Order approximately 65–145 million years ago. The most basal species in which the LGT was found is Plutella xylostella (superfamily: Yponomeutoidea). Array data from Bombyx mori shows that GH31 is expressed, and low dN/dS ratios indicates the LGT coding sequence is under strong stabilizing selection. These findings provide further support for the proposition that bacterial LGTs are relatively common in insects and likely to be an underappreciated source of adaptive genetic material. PMID:23533610

  2. The Atypical Inhibitor of NF-κB, IκBζ, Controls Macrophage Interleukin-10 Expression.

    PubMed

    Hörber, Sebastian; Hildebrand, Dominic G; Lieb, Wolfgang S; Lorscheid, Sebastian; Hailfinger, Stephan; Schulze-Osthoff, Klaus; Essmann, Frank

    2016-06-10

    Macrophages constitute a first line of pathogen defense by triggering a number of inflammatory responses and the secretion of various pro-inflammatory cytokines. Recently, we and others found that IκBζ, an atypical IκB family member and transcriptional coactivator of selected NF-κB target genes, is essential for macrophage expression of a subset of pro-inflammatory cytokines, such as IL-6, IL-12, and CCL2. Despite defective pro-inflammatory cytokine expression, however, IκBζ-deficient mice develop symptoms of chronic inflammation. To elucidate this discrepancy, we analyzed a regulatory role of IκBζ for the expression of anti-inflammatory cytokines and identified IκBζ as an essential activator of IL-10 expression. LPS-challenged peritoneal and bone marrow-derived macrophages from IκBζ-deficient mice revealed strongly decreased transcription and secretion of IL-10 compared with wild-type mice. Moreover, ectopic expression of IκBζ was sufficient to stimulate Il10 transcription. On the molecular level, IκBζ directly activated the Il10 promoter at a proximal κB site and was required for the transcription-enhancing trimethylation of histone 3 at lysine 4. Together, our findings show for the first time the IκBζ-dependent expression of an anti-inflammatory cytokine that is crucial in controlling immune responses. PMID:27129283

  3. Attenuation of methylglyoxal-induced peritoneal fibrosis: immunomodulation by interleukin-10.

    PubMed

    Onishi, Akira; Akimoto, Tetsu; Urabe, Masashi; Hirahara, Ichiro; Muto, Shigeaki; Ozawa, Keiya; Nagata, Daisuke; Kusano, Eiji

    2015-12-01

    Peritoneal fibrosis (PF), a serious pathophysiology of peritoneal dialysis (PD), is implicated in various types of chronic inflammation. In the present study, we examined the benefits of interleukin (IL)-10, which exerts anti-inflammatory effects, in an experimental rat model of methylglyoxal (MGO)-induced PF. We injected an adeno-associated virus (AAV) vector encoding rat IL-10 or enhanced green fluorescent protein (GFP) into male Sprague-Dawley rats at 6 weeks of age. Four weeks later, the rats received continuous peritoneal injections of conventional PD fluid (PDF) with MGO for 3 weeks. Then, the peritoneal histology and the expression levels of fibrogenic mediators and proinflammatory cytokines were analyzed. The rats demonstrating persistent IL-10 expression showed significantly reduced fibrous peritoneal thickening compared with those with GFP expression. The infiltration of macrophages, the expression of tumor necrosis factor-α, IL-1β, IL-6, transforming growth factor-β1, Snail, and matrix metalloproteinase 2 genes as well as the proliferation of mesenchymal-like mesothelial cells augmented by MGO were all significantly suppressed by IL-10 expression. IL-10 also abrogated the extent of MGO-induced bowel adhesions mimicking a cocoon-like mass. Our findings provide valuable insight into the potential benefit of immunomodulation with IL-10 as one potentially effective therapeutic strategy for preventing the onset of peritoneal injury resulting in PF. PMID:26367488

  4. Adenovirus serotype 5 hexon mediates liver gene transfer.

    PubMed

    Waddington, Simon N; McVey, John H; Bhella, David; Parker, Alan L; Barker, Kristeen; Atoda, Hideko; Pink, Rebecca; Buckley, Suzanne M K; Greig, Jenny A; Denby, Laura; Custers, Jerome; Morita, Takashi; Francischetti, Ivo M B; Monteiro, Robson Q; Barouch, Dan H; van Rooijen, Nico; Napoli, Claudio; Havenga, Menzo J E; Nicklin, Stuart A; Baker, Andrew H

    2008-02-01

    Adenoviruses are used extensively as gene transfer agents, both experimentally and clinically. However, targeting of liver cells by adenoviruses compromises their potential efficacy. In cell culture, the adenovirus serotype 5 fiber protein engages the coxsackievirus and adenovirus receptor (CAR) to bind cells. Paradoxically, following intravascular delivery, CAR is not used for liver transduction, implicating alternate pathways. Recently, we demonstrated that coagulation factor (F)X directly binds adenovirus leading to liver infection. Here, we show that FX binds to the Ad5 hexon, not fiber, via an interaction between the FX Gla domain and hypervariable regions of the hexon surface. Binding occurs in multiple human adenovirus serotypes. Liver infection by the FX-Ad5 complex is mediated through a heparin-binding exosite in the FX serine protease domain. This study reveals an unanticipated function for hexon in mediating liver gene transfer in vivo. PMID:18267072

  5. Gene transfer from a parasitic flowering plant to a fern

    PubMed Central

    Davis, Charles C; Anderson, William R; Wurdack, Kenneth J

    2005-01-01

    The rattlesnake fern (Botrychium virginianum (L.) Sw.) is obligately mycotrophic and widely distributed across the northern hemisphere. Three mitochondrial gene regions place this species with other ferns in Ophioglossaceae, while two regions place it as a member of the largely parasitic angiosperm order Santalales (sandalwoods and mistletoes). These discordant phylogenetic placements suggest that part of the genome in B. virginianum was acquired by horizontal gene transfer (HGT), perhaps from root-parasitic Loranthaceae. These transgenes are restricted to B. virginianum and occur across the range of the species. Molecular and life-history traits indicate that the transfer preceded the global expansion of B. virginianum, and that the latter may have happened very rapidly. This is the first report of HGT from an angiosperm to a fern, through either direct parasitism or the mediation of interconnecting fungal symbionts. PMID:16191635

  6. A Rice Stowaway MITE for Gene Transfer in Yeast

    PubMed Central

    Fattash, Isam; Bhardwaj, Priyanka; Hui, Caleb; Yang, Guojun

    2013-01-01

    Miniature inverted repeat transposable elements (MITEs) lack protein coding capacity and often share very limited sequence similarity with potential autonomous elements. Their capability of efficient transposition and dramatic amplification led to the proposition that MITEs are an untapped rich source of materials for transposable element (TE) based genetic tools. To test the concept of using MITE sequence in gene transfer, a rice Stowaway MITE previously shown to excise efficiently in yeast was engineered to carry cargo genes (neo and gfp) for delivery into the budding yeast genome. Efficient excision of the cargo gene cassettes was observed even though the excision frequency generally decreases with the increase of the cargo sizes. Excised elements insert into new genomic loci efficiently, with about 65% of the obtained insertion sites located in genes. Elements at the primary insertion sites can be remobilized, frequently resulting in copy number increase of the element. Surprisingly, the orientation of a cargo gene (neo) on a construct bearing dual reporter genes (gfp and neo) was found to have a dramatic effect on transposition frequency. These results demonstrated the concept that MITE sequences can be useful in engineering genetic tools to deliver cargo genes into eukaryotic genomes. PMID:23704977

  7. Detection of homologous horizontal gene transfer in SNP data

    Energy Science and Technology Software Center (ESTSC)

    2012-07-23

    We study the detection of mutations, sequencing errors, and homologous horizontal gene transfers (HGT) in a set of closely related microbial genomes. We base the model on single nucleotide polymorphisms (SNP's) and break the genomes into blocks to handle the rearrangement problem. Then we apply a synamic programming algorithm to model whether changes within each block are likely a result of mutations, sequencing errors, or HGT.

  8. Nitric oxide modulates lipopolysaccharide-induced endothelial platelet endothelial cell adhesion molecule expression via interleukin-10.

    PubMed

    Hebeda, C B; Teixeira, S A; Tamura, E K; Muscará, M N; de Mello, S B V; Markus, R P; Farsky, S H P

    2011-08-01

    We have shown previously that nitric oxide (NO) controls platelet endothelial cell adhesion molecule (PECAM-1) expression on both neutrophils and endothelial cells under physiological conditions. Here, the molecular mechanism by which NO regulates lipopolysaccharide (LPS)-induced endothelial PECAM-1 expression and the role of interleukin (IL)-10 on this control was investigated. For this purpose, N-(G)-nitro-L-arginine methyl ester (L-NAME; 20 mg/kg/day for 14 days dissolved in drinking water) was used to inhibit both constitutive (cNOS) and inducible nitric oxide (iNOS) synthase activities in LPS-stimulated Wistar rats (5 mg/kg, intraperitoneally). This treatment resulted in reduced levels of serum NO. Under this condition, circulating levels of IL-10 was enhanced, secreted mainly by circulating lymphocytes, dependent on transcriptional activation, and endothelial PECAM-1 expression was reduced independently on reduced gene synthesis. The connection between NO, IL-10 and PECAM-1 expression was examined by incubating LPS-stimulated (1 µg/ml) cultured endothelial cells obtained from naive rats with supernatant of LPS-stimulated lymphocytes, which were obtained from blood of control or L-NAME-treated rats. Supernatant of LPS-stimulated lymphocytes obtained from L-NAME-treated rats, which contained higher levels of IL-10, reduced LPS-induced PECAM-1 expression by endothelial cells, and this reduction was reversed by adding the anti-IL-10 monoclonal antibody. Therefore, an association between NO, IL-10 and PECAM-1 was found and may represent a novel mechanism by which NO controls endothelial cell functions. PMID:21564091

  9. Selective Gene Transfer to the Retina Using Intravitreal Ultrasound Irradiation

    PubMed Central

    Sonoda, Shozo; Tachibana, Katsuro; Yamashita, Toshifumi; Shirasawa, Makoto; Terasaki, Hiroto; Uchino, Eisuke; Suzuki, Ryo; Maruyama, Kazuo; Sakamoto, Taiji

    2012-01-01

    This paper aims to evaluate the efficacy of intravitreal ultrasound (US) irradiation for green fluorescent protein (GFP) plasmid transfer into the rabbit retina using a miniature US transducer. Intravitreal US irradiation was performed by a slight modification of the transconjunctival sutureless vitrectomy system utilizing a small probe. After vitrectomy, the US probe was inserted through a scleral incision. A mixture of GFP plasmid (50 μL) and bubble liposomes (BLs; 50 μL) was injected into the vitreous cavity, and US was generated to the retina using a SonoPore 4000. The control group was not exposed to US. After 72 h, the gene-transfer efficiency was quantified by counting the number of GFP-positive cells. The retinas that received plasmid, BL, and US showed a significant increase in the number (average ± SEM) of GFP-positive cells (32 ± 4.9; n = 7; P < 0.01 ). No GFP-positive cells were observed in the control eyes (n = 7). Intravitreal retinal US irradiation can transfer the GFP plasmid into the retina without causing any apparent damage. This procedure could be used to transfer genes and drugs directly to the retina and therefore has potential therapeutic value. PMID:22518277

  10. Comparison of Efficacy of the Disease-Specific LOX1- and Constitutive Cytomegalovirus-Promoters in Expressing Interleukin 10 through Adeno-Associated Virus 2/8 Delivery in Atherosclerotic Mice

    PubMed Central

    Zhu, Hongqing; Cao, Maohua; Mirandola, Leonardo; Figueroa, Jose A.; Cobos, Everardo; Chiriva-Internati, Maurizio; Hermonat, Paul L.

    2014-01-01

    The development of gene therapy vectors for treating diseases of the cardiovascular system continues at a steady pace. Moreover, in the field of gene therapy the utility of “disease-specific promoters” has strong appeal. Many therapeutic genes, including transforming growth factor beta 1 or interleukin 10, are associated to adverse effects. The use of a disease-specific promoter might minimize toxicity. The lectin-like oxidized low density lipoprotein receptor 1 is a marker of cardiovascular disease and a potential therapeutic target. The lectin-like oxidized low density lipoprotein receptor 1 is known to be up-regulated early during disease onset in a number of cell types at the sites where the disease will be clinically evident. In this study an adeno-associated virus-2 DNA vector (AAV2) using the AAV8 capsid, and containing the full length The lectin-like oxidized low density lipoprotein receptor 1 promoter, was generated and assayed for its ability to express human interleukin 10 in low density lipoprotein receptor knockout mice on high cholesterol diet. The cytomegalovirus early promoter was used for comparison in a similarly structured vector. The two promoters were found to have equal efficacy in reducing atherogenesis as measured by aortic systolic blood velocity, aortic cross sectional area, and aortic wall thickness. This is the first head-to-head comparison of a constitutive with a disease-specific promoter in a therapeutic context. These data strongly suggest that the use of a disease-specific promoter is appropriate for therapeutic gene delivery. PMID:24736312

  11. Dynamic monitoring of horizontal gene transfer in soil

    NASA Astrophysics Data System (ADS)

    Cheng, H. Y.; Masiello, C. A.; Silberg, J. J.; Bennett, G. N.

    2015-12-01

    Soil microbial gene expression underlies microbial behaviors (phenotypes) central to many aspects of C, N, and H2O cycling. However, continuous monitoring of microbial gene expression in soils is challenging because genetically-encoded reporter proteins widely used in the lab are difficult to deploy in soil matrices: for example, green fluorescent protein cannot be easily visualized in soils, even in the lab. To address this problem we have developed a reporter protein that releases small volatile gases. Here, we applied this gas reporter in a proof-of-concept soil experiment, monitoring horizontal gene transfer, a microbial activity that alters microbial genotypes and phenotypes. Horizontal gene transfer is central to bacterial evolution and adaptation and is relevant to problems such as the spread of antibiotic resistance, increasing metal tolerance in superfund sites, and bioremediation capability of bacterial consortia. This process is likely to be impacted by a number of matrix properties not well-represented in the petri dish, such as microscale variations in water, nutrients, and O2, making petri-dish experiments a poor proxy for environmental processes. We built a conjugation system using synthetic biology to demonstrate the use of gas-reporting biosensors in safe, lab-based biogeochemistry experiments, and here we report the use of these sensors to monitor horizontal gene transfer in soils. Our system is based on the F-plasmid conjugation in Escherichia coli. We have found that the gas signal reports on the number of cells that acquire F-plasmids (transconjugants) in a loamy Alfisol collected from Kellogg Biological Station. We will report how a gas signal generated by transconjugants varies with the number of F-plasmid donor and acceptor cells seeded in a soil, soil moisture, and soil O2 levels.

  12. [Synthesis of new gene-loaded microbubbles serve as gene delivery vehicle applied in reporter gene transfer into cardiac myocytes].

    PubMed

    Wang, Guozhong; Hu, Shenjiang; Zheng, Zhelan; Sun, Jian; Zheng, Xia; Zhu, Zhaohui; Li, Jiang; Yao, Yumei

    2006-08-01

    To improve the stability and gene-carried capability of gene-attached microbubbles, the method for manufacture of albumin microbubbles was modified and new gene-loaded microbubbles were synthesized by incorporated gene-PEI complex into the shell of microbubbles. Agarose gel electrophoresis and bacteria transformation showed that PEI had the ability to provide the protection of plasmid DNA from ultrasonic degradation. The new gene-loaded microbubbles exhibited excellent acoustical and hemorheological properties. Moreover, they could carry more plasmid DNA than gene-attached microbubbles. beta-galactosidase plasmid transfection into cardiac myocytes was performed by using ultrasound targeted destruction of new gene-loaded microbubbles or gene-attached microbubbles. Gene expression in cardiac myocytes was detected by beta-galactosidase in situ staining and quantitive assay. It was shown that beta-galactosidase activity in cardiac myocytes was enhanced 107-fold by ultrasonic destruction of gene-loaded microbubbles compared with naked plasmid transfection and new gene-loaded microbubbles resulted in 6.85-fold increase in beta-galactosidase activity compared with optimal transfection mediated by gene-attached microbubbles. These results suggested that ultrasonic destruction of the gene-loaded microbubbles can enhance the cardiac myocytes exogenous gene transfer efficiency significantly and new gene-loaded microbubbles is an efficient and safe gene delivery vehicle. PMID:17002125

  13. Horizontal Gene Transfer Contributes to the Evolution of Arthropod Herbivory

    PubMed Central

    Wybouw, Nicky; Pauchet, Yannick; Heckel, David G.; Van Leeuwen, Thomas

    2016-01-01

    Within animals, evolutionary transition toward herbivory is severely limited by the hostile characteristics of plants. Arthropods have nonetheless counteracted many nutritional and defensive barriers imposed by plants and are currently considered as the most successful animal herbivores in terrestrial ecosystems. We gather a body of evidence showing that genomes of various plant feeding insects and mites possess genes whose presence can only be explained by horizontal gene transfer (HGT). HGT is the asexual transmission of genetic information between reproductively isolated species. Although HGT is known to have great adaptive significance in prokaryotes, its impact on eukaryotic evolution remains obscure. Here, we show that laterally transferred genes into arthropods underpin many adaptations to phytophagy, including efficient assimilation and detoxification of plant produced metabolites. Horizontally acquired genes and the traits they encode often functionally diversify within arthropod recipients, enabling the colonization of more host plant species and organs. We demonstrate that HGT can drive metazoan evolution by uncovering its prominent role in the adaptations of arthropods to exploit plants. PMID:27307274

  14. Evidence of horizontal gene transfer between obligate leaf nodule symbionts.

    PubMed

    Pinto-Carbó, Marta; Sieber, Simon; Dessein, Steven; Wicker, Thomas; Verstraete, Brecht; Gademann, Karl; Eberl, Leo; Carlier, Aurelien

    2016-09-01

    Bacteria of the genus Burkholderia establish an obligate symbiosis with plant species of the Rubiaceae and Primulaceae families. The bacteria, housed within the leaves, are transmitted hereditarily and have not yet been cultured. We have sequenced and compared the genomes of eight bacterial leaf nodule symbionts of the Rubiaceae plant family. All of the genomes exhibit features consistent with genome erosion. Genes potentially involved in the biosynthesis of kirkamide, an insecticidal C7N aminocyclitol, are conserved in most Rubiaceae symbionts. However, some have partially lost the kirkamide pathway due to genome erosion and are unable to synthesize the compound. Kirkamide synthesis is therefore not responsible for the obligate nature of the symbiosis. More importantly, we find evidence of intra-clade horizontal gene transfer (HGT) events affecting genes of the secondary metabolism. This indicates that substantial gene flow can occur at the early stages following host restriction in leaf nodule symbioses. We propose that host-switching events and plasmid conjugative transfers could have promoted these HGTs. This genomic analysis of leaf nodule symbionts gives, for the first time, new insights in the genome evolution of obligate symbionts in their early stages of the association with plants. PMID:26978165

  15. Horizontal Gene Transfer Contributes to the Evolution of Arthropod Herbivory.

    PubMed

    Wybouw, Nicky; Pauchet, Yannick; Heckel, David G; Van Leeuwen, Thomas

    2016-01-01

    Within animals, evolutionary transition toward herbivory is severely limited by the hostile characteristics of plants. Arthropods have nonetheless counteracted many nutritional and defensive barriers imposed by plants and are currently considered as the most successful animal herbivores in terrestrial ecosystems. We gather a body of evidence showing that genomes of various plant feeding insects and mites possess genes whose presence can only be explained by horizontal gene transfer (HGT). HGT is the asexual transmission of genetic information between reproductively isolated species. Although HGT is known to have great adaptive significance in prokaryotes, its impact on eukaryotic evolution remains obscure. Here, we show that laterally transferred genes into arthropods underpin many adaptations to phytophagy, including efficient assimilation and detoxification of plant produced metabolites. Horizontally acquired genes and the traits they encode often functionally diversify within arthropod recipients, enabling the colonization of more host plant species and organs. We demonstrate that HGT can drive metazoan evolution by uncovering its prominent role in the adaptations of arthropods to exploit plants. PMID:27307274

  16. PEGylated Cationic Serum Albumin for Boosting Retroviral Gene Transfer.

    PubMed

    Palesch, David; Boldt, Felix; Müller, Janis A; Eisele, Klaus; Stürzel, Christina M; Wu, Yuzhou; Münch, Jan; Weil, Tanja

    2016-08-17

    Retroviral vectors are common tools for introducing genes into the genome of a cell. However, low transduction rates are a major limitation in retroviral gene transfer, especially in clinical applications. We generated cationic human serum albumin (cHSA) protected by a shell of poly(ethylene glycol) (PEG); this significantly enhanced retroviral gene transduction with potentially attractive pharmacokinetics and low immunogenicity. By screening a panel of chemically optimized HSA compounds, we identified a very potent enhancer that boosted the transduction rates of viral vectors. Confocal microscopy revealed a drastically increased number of viral particles attached to the surfaces of target cells. In accordance with the positive net charge of cationic and PEGylated HSA, this suggests a mechanism of action in which the repulsion of the negatively charged cellular and viral vector membranes is neutralized, thereby promoting attachment and ultimately transduction. Importantly, the transduction-enhancing PEGylated HSA derivative evaded recognition by HSA-specific antibodies and macrophage activation. Our findings hold great promise for facilitating improved retroviral gene transfer. PMID:27239020

  17. HGTree: database of horizontally transferred genes determined by tree reconciliation

    PubMed Central

    Jeong, Hyeonsoo; Sung, Samsun; Kwon, Taehyung; Seo, Minseok; Caetano-Anollés, Kelsey; Choi, Sang Ho; Cho, Seoae; Nasir, Arshan; Kim, Heebal

    2016-01-01

    The HGTree database provides putative genome-wide horizontal gene transfer (HGT) information for 2472 completely sequenced prokaryotic genomes. This task is accomplished by reconstructing approximate maximum likelihood phylogenetic trees for each orthologous gene and corresponding 16S rRNA reference species sets and then reconciling the two trees under parsimony framework. The tree reconciliation method is generally considered to be a reliable way to detect HGT events but its practical use has remained limited because the method is computationally intensive and conceptually challenging. In this regard, HGTree (http://hgtree.snu.ac.kr) represents a useful addition to the biological community and enables quick and easy retrieval of information for HGT-acquired genes to better understand microbial taxonomy and evolution. The database is freely available and can be easily scaled and updated to keep pace with the rapid rise in genomic information. PMID:26578597

  18. PCR-based detection of gene transfer vectors: application to gene doping surveillance.

    PubMed

    Perez, Irene C; Le Guiner, Caroline; Ni, Weiyi; Lyles, Jennifer; Moullier, Philippe; Snyder, Richard O

    2013-12-01

    Athletes who illicitly use drugs to enhance their athletic performance are at risk of being banned from sports competitions. Consequently, some athletes may seek new doping methods that they expect to be capable of circumventing detection. With advances in gene transfer vector design and therapeutic gene transfer, and demonstrations of safety and therapeutic benefit in humans, there is an increased probability of the pursuit of gene doping by athletes. In anticipation of the potential for gene doping, assays have been established to directly detect complementary DNA of genes that are top candidates for use in doping, as well as vector control elements. The development of molecular assays that are capable of exposing gene doping in sports can serve as a deterrent and may also identify athletes who have illicitly used gene transfer for performance enhancement. PCR-based methods to detect foreign DNA with high reliability, sensitivity, and specificity include TaqMan real-time PCR, nested PCR, and internal threshold control PCR. PMID:23912835

  19. Indications for Acquisition of Reductive Dehalogenase Genes through Horizontal Gene Transfer by Dehalococcoides ethenogenes Strain 195

    PubMed Central

    Regeard, Christophe; Maillard, Julien; Dufraigne, Christine; Deschavanne, Patrick; Holliger, Christof

    2005-01-01

    The genome of Dehalococcoides ethenogenes strain 195, an anaerobic dehalorespiring bacterium, contains 18 copies of putative reductive dehalogenase genes, including the well-characterized tceA gene, whose gene product functions as the key enzyme in the environmentally important dehalorespiration process. The genome of D. ethenogenes was analyzed using a bioinformatic tool based on the frequency of oligonucleotides. The results in the form of a genomic signature revealed several local disruptions of the host signature along the genome sequence. These fractures represent DNA segments of potentially foreign origin, so-called atypical regions, which may have been acquired by an ancestor through horizontal gene transfer. Most interestingly, 15 of the 18 reductive dehalogenase genes, including the tceA gene, were found to be located in these regions, strongly indicating the foreign nature of the dehalorespiration activity. The GC content and the presence of recombinase genes within some of these regions corroborate this hypothesis. A hierarchical classification of the atypical regions containing the reductive dehalogenase genes indicated that these regions were probably acquired by several gene transfer events. PMID:15932990

  20. Effects of recombinant human interleukin-10 on Treg cells, IL-10 and TGF-β in transplantation of rabbit skin.

    PubMed

    Liu, Kai Shan; Fan, Xiao Qin; Zhang, Lei; Wen, Qiong Na; Feng, Ji Hong; Chen, Fu Chao; Luo, Jun Min; Sun, Wan Bang

    2014-02-01

    The current study aimed to investigate the rejection and survival time of grafted skin, and the changes of Treg cells, interleukin 10 (IL-10) and transforming growth factor-β (TGF-β) in peripheral blood following skin transplantation with recombinant human interleukin-10 (rhIL-10) or cyclosporin A (CsA), as well as the role of IL-10 in immunological rejection mechanisms. A total of 36 rabbits were divided into two groups. The skin of a donor rabbit was transplanted onto the back of one receptor rabbit. Receptors were randomly divided into six groups, including rhIL-10 low-dose (5 µg/kg/d), rhIL-10 high-dose (10 µg/kg/d), CsA low-dose (5 mg/kg/d), CsA high-dose (10 mg/kg/d), rhIL-10 (5 µg/kg/d) and CsA (5 mg/kg/d) and negative control normal saline (NS; 1 ml/d). All groups received intramuscular drug injection for ten days, beginning one day prior to skin transplantation surgery. Following transplantation, each rabbit's peripheral blood was collected at different times. The changes of CD4+CD25+ regulatory T cells, IL-10 and TGF-β were determined by flow cytometry and enzyme-linked immunosorbent assay. When compared with the control group, the rejection and survival times of the experimental groups were longer following skin graft. Compared with the two CsA groups and the control group, the proportion of CD4+CD25+ regulatory T cells of rhIL-10 groups was significantly upregulated on the 4th and 7th days following surgery. However, TGF-β levels were not significantly different. Data suggested that the concentration of IL-10 was positively correlated with the proportion of CD4+CD25+ regulatory T cells. In addition, IL-10 may delay the rejection time of rabbit skin transplantation and prolong the survival time. Thus, the role of IL-10 in inhibited allograft rejection may be associated with CD4+CD25+ regulatory T cells and IL-10, and may be independent of TGF-β. PMID:24270972

  1. Bacterial gene transfer by natural genetic transformation in the environment.

    PubMed Central

    Lorenz, M G; Wackernagel, W

    1994-01-01

    Natural genetic transformation is the active uptake of free DNA by bacterial cells and the heritable incorporation of its genetic information. Since the famous discovery of transformation in Streptococcus pneumoniae by Griffith in 1928 and the demonstration of DNA as the transforming principle by Avery and coworkers in 1944, cellular processes involved in transformation have been studied extensively by in vitro experimentation with a few transformable species. Only more recently has it been considered that transformation may be a powerful mechanism of horizontal gene transfer in natural bacterial populations. In this review the current understanding of the biology of transformation is summarized to provide the platform on which aspects of bacterial transformation in water, soil, and sediments and the habitat of pathogens are discussed. Direct and indirect evidence for gene transfer routes by transformation within species and between different species will be presented, along with data suggesting that plasmids as well as chromosomal DNA are subject to genetic exchange via transformation. Experiments exploring the prerequisites for transformation in the environment, including the production and persistence of free DNA and factors important for the uptake of DNA by cells, will be compiled, as well as possible natural barriers to transformation. The efficiency of gene transfer by transformation in bacterial habitats is possibly genetically adjusted to submaximal levels. The fact that natural transformation has been detected among bacteria from all trophic and taxonomic groups including archaebacteria suggests that transformability evolved early in phylogeny. Probable functions of DNA uptake other than gene acquisition will be discussed. The body of information presently available suggests that transformation has a great impact on bacterial population dynamics as well as on bacterial evolution and speciation. PMID:7968924

  2. In vivo Cytokine Gene Transfer by Gene Gun Reduces Tumor Growth in Mice

    NASA Astrophysics Data System (ADS)

    Sun, Wenn H.; Burkholder, Joseph K.; Sun, Jian; Culp, Jerilyn; Turner, Joel; Lu, Xing G.; Pugh, Thomas D.; Ershler, William B.; Yang, Ning-Sun

    1995-03-01

    Implantation of tumor cells modified by in vitro cytokine gene transfer has been shown by many investigators to result in potent in vivo antitumor activities in mice. Here we describe an approach to tumor immunotherapy utilizing direct transfection of cytokine genes into tumorbearing animals by particle-mediated gene transfer. In vivo transfection of the human interleukin 6 gene into the tumor site reduced methylcholanthrene-induced fibrosarcoma growth, and a combination of murine tumor necrosis factor α and interferon γ genes inhibited growth of a renal carcinoma tumor model (Renca). In addition, treatment with murine interleukin 2 and interferon γ genes prolonged the survival of Renca tumor-bearing mice and resulted in tumor eradication in 25% of the test animals. Transgene expression was demonstrated in treated tissues by ELISA and immunohistochemical analysis. Significant serum levels of interleukin 6 and interferon γ were detected, demonstrating effective secretion of transgenic proteins from treated skin into the bloodstream. This in vivo cytokine gene therapy approach provides a system for evaluating the antitumor properties of various cytokines in different tumor models and has potential utility for human cancer gene therapy.

  3. Intrathecal synthesis of interleukin-10 (IL-10) in viral and inflammatory diseases of the central nervous system.

    PubMed

    Gallo, P; Sivieri, S; Rinaldi, L; Yan, X B; Lolli, F; De Rossi, A; Tavolato, B

    1994-10-01

    The intrathecal synthesis of interleukin 10 (IL-10) was investigated in 120 paired cerebrospinal fluid (CSF) and serum specimens from patients with various inflammatory and non-inflammatory diseases of the central nervous system (CNS). IL-10 was not demonstrated in the sera, but detectable levels were found in the CSF from: patients with acute viral ("aseptic") meningitis, but only within 48-72 h of symptom onset; human immunodeficiency virus type 1 (HIV)-infected patients with HIV-related encephalitis/leukoencephalopathy or cryptococcal meningitis; a patient with primary B cell lymphoma of the CNS, and a patient with encephalomeningeal sarcoidosis (in whom IL-10 was demonstrated in all CSF collected over a period of 6-months). In chronic meningeal infections/inflammations, IL-10 seems to be continuously produced within the CSF. Our findings suggest that IL-10, a cytokine which exerts many immunosuppressive actions, may play different immunomodulatory roles in CNS diseases; in particular, its intrathecal synthesis may explain why some infectious and inflammatory meningeal diseases may have slow development and chronic evolution. PMID:7836946

  4. Anti-inflammatory pathways and alcoholic liver disease: role of an adiponectin/interleukin-10/heme oxygenase-1 pathway.

    PubMed

    Mandal, Palash; Pritchard, Michele T; Nagy, Laura E

    2010-03-21

    The development of alcoholic liver disease (ALD) is a complex process involving both the parenchymal and non-parenchymal cells in the liver. Enhanced inflammation in the liver during ethanol exposure is an important contributor to injury. Kupffer cells, the resident macrophages in liver, are particularly critical to the onset of ethanol-induced liver injury. Chronic ethanol exposure sensitizes Kupffer cells to activation by lipopolysaccharide via Toll-like receptor 4. This sensitization enhances production of inflammatory mediators, such as tumor necrosis factor-alpha and reactive oxygen species, that contribute to hepatocyte dysfunction, necrosis, apoptosis, and fibrosis. Impaired resolution of the inflammatory process probably also contributes to ALD. The resolution of inflammation is an active, highly coordinated response that can potentially be manipulated via therapeutic interventions to treat chronic inflammatory diseases. Recent studies have identified an adiponectin/interleukin-10/heme oxygenase-1 (HO-1) pathway that is profoundly effective in dampening the enhanced activation of innate immune responses in primary cultures of Kupffer cells, as well as in an in vivo mouse model of chronic ethanol feeding. Importantly, induction of HO-1 also reduces ethanol-induced hepatocellular apoptosis in this in vivo model. Based on these data, we hypothesize that the development of therapeutic agents to regulate HO-1 and its downstream targets could be useful in enhancing the resolution of inflammation during ALD and preventing progression of early stages of liver injury. PMID:20238399

  5. A Defect in Interleukin-10 Leads to Enhanced Malarial Disease in Plasmodium chabaudi chabaudi Infection in Mice

    PubMed Central

    Li, Ching; Corraliza, Inés; Langhorne, Jean

    1999-01-01

    Infection of interleukin-10 (IL-10)-nonexpressing (IL-10−/−) mice with Plasmodium chabaudi chabaudi (AS) leads to exacerbated pathology in female mice and death in a proportion of them. Hypoglycemia, hypothermia, and loss in body weight were significantly greater in female IL-10−/− mice than in male knockout mice and all wild-type (WT) mice during the acute phase of infection. At this time, both female and male IL-10−/− mice produced more gamma interferon (IFN-γ), tumor necrosis factor alpha (TNF-α), and IL-12p40 mRNA than their respective WT counterparts. Inactivation of IFN-γ in IL-10−/− mice by the injection of anti-IFN-γ antibodies or by the generation of IL-10−/− IFN-γ receptor−/− double-knockout mice resulted in reduced mortality but did not affect body weight, temperature, or blood glucose levels. The data suggest that IFN-γ-independent pathways may be responsible for these pathological features of P. chabaudi malaria and may be due to direct stimulation of TNF-α by the parasite. Since male and female knockout mice both produce more inflammatory cytokines than their WT counterparts, it is likely that the mortality seen in females is due to the nature or magnitude of the response to these cytokines rather than the amount of IFN-γ or TNF-α produced. PMID:10456884

  6. Interleukin-10 plays a key role in the modulation of neutrophils recruitment and lung inflammation during infection by Streptococcus pneumoniae

    PubMed Central

    Peñaloza, Hernán F; Nieto, Pamela A; Muñoz-Durango, Natalia; Salazar-Echegarai, Francisco J; Torres, Javiera; Parga, María J; Alvarez-Lobos, Manuel; Riedel, Claudia A; Kalergis, Alexis M; Bueno, Susan M

    2015-01-01

    Streptococcus pneumoniae is a major aetiological agent of pneumonia worldwide, as well as otitis media, sinusitis, meningitis and sepsis. Recent reports have suggested that inflammation of lungs due to S. pneumoniae infection promotes bacterial dissemination and severe disease. However, the contribution of anti-inflammatory molecules to the pathogenesis of S. pneumoniae remains unknown. To elucidate whether the production of the anti-inflammatory cytokine interleukin-10 (IL-10) is beneficial or detrimental for the host during pneumococcal pneumonia, we performed S. pneumoniae infections in mice lacking IL-10 (IL-10−/− mice). The IL-10−/− mice showed increased mortality, higher expression of pro-inflammatory cytokines, and an exacerbated recruitment of neutrophils into the lungs after S. pneumoniae infection. However, IL-10−/− mice showed significantly lower bacterial loads in lungs, spleen, brain and blood, when compared with mice that produced this cytokine. Our results support the notion that production of IL-10 during S. pneumoniae infection modulates the expression of pro-inflammatory cytokines and the infiltration of neutrophils into the lungs. This feature of IL-10 is important to avoid excessive inflammation of tissues and to improve host survival, even though bacterial dissemination is less efficient in the absence of this cytokine. PMID:26032199

  7. Differential Induction of Interleukin-10 in Monocytes by HIV-1 Clade B and Clade C Tat Proteins*

    PubMed Central

    Wong, Justine K.; Campbell, Grant R.; Spector, Stephen A.

    2010-01-01

    The clade B human immunodeficiency virus, type 1 (HIV-1) Tat (trans-acting regulatory protein) induces interleukin-10 (IL-10) production in monocytes. IL-10, an anti-inflammatory cytokine, down-regulates proinflammatory cytokines and suppresses the immune response, leading to a rapid progression from HIV-1 infection to AIDS. Nine clades of HIV-1 are responsible for the majority of infections worldwide. Recent studies demonstrate that different HIV-1 clades have biological differences in relation to transmission, replication, and disease progression. In this study, we show that the cysteine to serine mutation at position 31, found in >90% of HIV-1 clade C Tat proteins, results in a marked decrease in IL-10 production in monocytes compared with clade B Tat. Additionally, the C31S mutation found in C Tat is responsible for the inability of these Tat proteins to produce high IL-10 levels in monocytes due to its inability to induce intracellular calcium flux through L-type calcium channels. Moreover, we show that p38α/p38β and phosphoinositide 3-kinase are crucial to Tat-induced IL-10 production. These findings provide further evidence that HIV-1 clades differ in their biological properties that may impact HIV-1 pathogenesis and disease progression. PMID:20378550

  8. Resveratrol induces the expression of interleukin-10 and brain-derived neurotrophic factor in BV2 microglia under hypoxia.

    PubMed

    Song, Juhyun; Cheon, So Yeong; Jung, Wonsug; Lee, Won Taek; Lee, Jong Eun

    2014-01-01

    Microglia are the resident macrophages of the central nervous system (CNS) and play an important role in neuronal recovery by scavenging damaged neurons. However, overactivation of microglia leads to neuronal death that is associated with CNS disorders. Therefore, regulation of microglial activation has been suggested to be an important target for treatment of CNS diseases. In the present study, we investigated the beneficial effect of resveratrol, a natural phenol with antioxidant effects, in the microglial cell line, BV2, in a model of hypoxia injury. Resveratrol suppressed the mRNA expression of the pro-inflammatory molecule, tumor necrosis factor-α, and promoted the mRNA expression of the anti-inflammatory molecule, interleukin-10, in BV2 microglia under hypoxic conditions. In addition, resveratrol inhibited the activation of the transcription factor, nuclear factor kappa-light-chain enhancer of activated B cells (NF-κB), which is upstream in the control of inflammatory reactions in hypoxia-injured BV2 microglia. Moreover, resveratrol promoted the expression of brain-derived neurotrophic factor (BDNF) in BV2 microglia under hypoxic stress. Overall, resveratrol may promote the beneficial function of microglia in ischemic brain injury. PMID:25184950

  9. Interleukin-10 plays a key role in the modulation of neutrophils recruitment and lung inflammation during infection by Streptococcus pneumoniae.

    PubMed

    Peñaloza, Hernán F; Nieto, Pamela A; Muñoz-Durango, Natalia; Salazar-Echegarai, Francisco J; Torres, Javiera; Parga, María J; Alvarez-Lobos, Manuel; Riedel, Claudia A; Kalergis, Alexis M; Bueno, Susan M

    2015-09-01

    Streptococcus pneumoniae is a major aetiological agent of pneumonia worldwide, as well as otitis media, sinusitis, meningitis and sepsis. Recent reports have suggested that inflammation of lungs due to S. pneumoniae infection promotes bacterial dissemination and severe disease. However, the contribution of anti-inflammatory molecules to the pathogenesis of S. pneumoniae remains unknown. To elucidate whether the production of the anti-inflammatory cytokine interleukin-10 (IL-10) is beneficial or detrimental for the host during pneumococcal pneumonia, we performed S. pneumoniae infections in mice lacking IL-10 (IL-10(-/-) mice). The IL-10(-/-) mice showed increased mortality, higher expression of pro-inflammatory cytokines, and an exacerbated recruitment of neutrophils into the lungs after S. pneumoniae infection. However, IL-10(-/-) mice showed significantly lower bacterial loads in lungs, spleen, brain and blood, when compared with mice that produced this cytokine. Our results support the notion that production of IL-10 during S. pneumoniae infection modulates the expression of pro-inflammatory cytokines and the infiltration of neutrophils into the lungs. This feature of IL-10 is important to avoid excessive inflammation of tissues and to improve host survival, even though bacterial dissemination is less efficient in the absence of this cytokine. PMID:26032199

  10. Adaptive Horizontal Gene Transfers between Multiple Cheese-Associated Fungi.

    PubMed

    Ropars, Jeanne; Rodríguez de la Vega, Ricardo C; López-Villavicencio, Manuela; Gouzy, Jérôme; Sallet, Erika; Dumas, Émilie; Lacoste, Sandrine; Debuchy, Robert; Dupont, Joëlle; Branca, Antoine; Giraud, Tatiana

    2015-10-01

    Domestication is an excellent model for studies of adaptation because it involves recent and strong selection on a few, identified traits [1-5]. Few studies have focused on the domestication of fungi, with notable exceptions [6-11], despite their importance to bioindustry [12] and to a general understanding of adaptation in eukaryotes [5]. Penicillium fungi are ubiquitous molds among which two distantly related species have been independently selected for cheese making-P. roqueforti for blue cheeses like Roquefort and P. camemberti for soft cheeses like Camembert. The selected traits include morphology, aromatic profile, lipolytic and proteolytic activities, and ability to grow at low temperatures, in a matrix containing bacterial and fungal competitors [13-15]. By comparing the genomes of ten Penicillium species, we show that adaptation to cheese was associated with multiple recent horizontal transfers of large genomic regions carrying crucial metabolic genes. We identified seven horizontally transferred regions (HTRs) spanning more than 10 kb each, flanked by specific transposable elements, and displaying nearly 100% identity between distant Penicillium species. Two HTRs carried genes with functions involved in the utilization of cheese nutrients or competition and were found nearly identical in multiple strains and species of cheese-associated Penicillium fungi, indicating recent selective sweeps; they were experimentally associated with faster growth and greater competitiveness on cheese and contained genes highly expressed in the early stage of cheese maturation. These findings have industrial and food safety implications and improve our understanding of the processes of adaptation to rapid environmental changes. PMID:26412136

  11. African vs. Caucasian and Asian difference for the association of interleukin-10 promotor polymorphisms with type 2 diabetes mellitus (a meta-analysis study)

    PubMed Central

    Tarabay, Mohammad; Elshazli, Rami; Settin, Ahmad

    2016-01-01

    Background Interleukin-10 (IL-10) is a multifunctional regulatory cytokine that might be associated with increased risk of type 2 diabetes mellitus (T2DM). IL-10 gene polymorphisms have been reported to be associated with T2DM in several ethnic populations with controversial results. Objectives This work is an updated meta-analysis aiming at the evaluation of the association between IL-10 gene polymorphisms: rs1800872 (− 592 C > A), rs1800896 (− 1082 A > G) and rs1800871 (− 819 C > T) with the risk of T2DM. Methods All available full text studies published up to July 2015 were included in this meta-analysis. Mainly Pubmed and Science Direct databases were searched for all eligible studies pertinent to testing the association between IL-10 gene polymorphisms with the susceptibility to T2DM. Further analyses of the pooled and stratified data in terms of individual polymorphic types and subject ethnicity were done and assessed using varied genetic models. Results Fifteen case-control studies with a total of 26 comparisons (10 for IL-10 − 592 C > A rs1800872, 11 for IL-10 − 1082 A > G rs1800896 and 5 for IL-10 − 819 C > T rs1800871 polymorphisms) met our inclusion criteria. IL-10 − 1082 A > G polymorphism was the only one to show an association with T2DM in all pooled sample particularly among Asian and European (high frequency of the G allele) ethnic groups. On the other hand, IL-10 − 592 C > A and − 819 C > T were significantly associated with T2DM only among African subjects. Conclusions This meta-analysis demonstrated that IL-10 − 1082 A > G polymorphism was associated with increased risk of development of T2DM in total subjects no matter was their ethnic background, while both IL-10 − 592 C > A and − 819 C > T polymorphisms were associated with that risk only among African subjects. PMID:27114918

  12. Interaction between Conjugative and Retrotransposable Elements in Horizontal Gene Transfer

    PubMed Central

    Novikova, Olga; Smith, Dorie; Hahn, Ingrid; Beauregard, Arthur; Belfort, Marlene

    2014-01-01

    Mobile genetic elements either encode their own mobilization machineries or hijack them from other mobile elements. Multiple classes of mobile elements often coexist within genomes and it is unclear whether they have the capacity to functionally interact and even collaborate. We investigate the possibility that molecular machineries of disparate mobile elements may functionally interact, using the example of a retrotransposon, in the form of a mobile group II intron, found on a conjugative plasmid pRS01 in Lactococcus lactis. This intron resides within the pRS01 ltrB gene encoding relaxase, the enzyme required for nicking the transfer origin (oriT) for conjugal transmission of the plasmid into a recipient cell. Here, we show that relaxase stimulates both the frequency and diversity of retrotransposition events using a retromobility indicator gene (RIG), and by developing a high-throughput genomic retrotransposition detection system called RIG-Seq. We demonstrate that LtrB relaxase not only nicks ssDNA of its cognate oriT in a sequence- and strand-specific manner, but also possesses weak off-target activity. Together, the data support a model in which the two different mobile elements, one using an RNA-based mechanism, the other using DNA-based transfer, do functionally interact. Intron splicing facilitates relaxase expression required for conjugation, whereas relaxase introduces spurious nicks in recipient DNA that stimulate both the frequency of intron mobility and the density of events. We hypothesize that this functional interaction between the mobile elements would promote horizontal conjugal gene transfer while stimulating intron dissemination in the donor and recipient cells. PMID:25474706

  13. Gene Transfer and Molecular Cloning of the Human NGF Receptor

    NASA Astrophysics Data System (ADS)

    Chao, Moses V.; Bothwell, Mark A.; Ross, Alonzo H.; Koprowski, Hilary; Lanahan, Anthony A.; Buck, C. Randall; Sehgal, Amita

    1986-04-01

    Nerve growth factor (NGF) and its receptor are important in the development of cells derived from the neural crest. Mouse L cell transformants have been generated that stably express the human NGF receptor gene transfer with total human DNA. Affinity cross-linking, metabolic labeling and immunoprecipitation, and equilibrium binding with 125I-labeled NGF revealed that this NGF receptor had the same size and binding characteristics as the receptor from human melanoma cells and rat PC12 cells. The sequences encoding the NGF receptor were molecularly cloned using the human Alu repetitive sequence as a probe. A cosmid clone that contained the human NGF receptor gene allowed efficient transfection and expression of the receptor.

  14. Site-Specific Gene Expression in Vivo by Direct Gene Transfer into the Arterial Wall

    NASA Astrophysics Data System (ADS)

    Nabel, Elizabeth G.; Plautz, Gregory; Nabel, Gary J.

    1990-09-01

    A recombinant β-galactosidase gene has been expressed in a specific arterial segment in vivo by direct infection with a murine amphotropic retroviral vector or by DNA transfection with the use of liposomes. Several cell types in the vessel wall were transduced, including endothelial and vascular smooth muscle cells. After retroviral infection, a recombinant reporter gene was expressed for at least 5 months, and no helper virus was detected. Recombinant gene expression achieved by direct retroviral infection or liposome-mediated DNA transfection was limited to the site of infection and was absent from liver, lung, kidney, and spleen. These results demonstrate that site-specific gene expression can be achieved by direct gene transfer in vivo and could be applied to the treatment of such human diseases as atherosclerosis or cancer.

  15. Laterally Transferred Gene Recruited as a Venom in Parasitoid Wasps.

    PubMed

    Martinson, Ellen O; Martinson, Vincent G; Edwards, Rachel; Mrinalini; Werren, John H

    2016-04-01

    Parasitoid wasps use venom to manipulate the immunity and metabolism of their host insects in a variety of ways to provide resources for their offspring. Yet, how genes are recruited and evolve to perform venom functions remain open questions. A recently recognized source of eukaryotic genome innovation is lateral gene transfer (LGT). Glycoside hydrolase family 19 (GH19) chitinases are widespread in bacteria, microsporidia, and plants where they are used in nutrient acquisition or defense, but have previously not been known in metazoans. In this study, a GH19 chitinase LGT is described from the unicellular microsporidia/Rozella clade into parasitoid wasps of the superfamily Chalcidoidea, where it has become recruited as a venom protein. The GH19 chitinase is present in 15 species of chalcidoid wasps representing four families, and phylogenetic analysis indicates that it was laterally transferred near or before the origin of Chalcidoidea (∼95 Ma). The GH19 chitinase gene is highly expressed in the venom gland of at least seven species, indicating a role in the complex host manipulations performed by parasitoid wasp venom. RNAi knockdown in the model parasitoid Nasonia vitripennis reveals that-following envenomation-the GH19 chitinase induces fly hosts to upregulate genes involved in an immune response to fungi. A second, independent LGT of GH19 chitinase from microsporidia into mosquitoes was also found, also supported by phylogenetic reconstructions. Besides these two LGT events, GH19 chitinase is not found in any other sequenced animal genome, or in any fungi outside the microsporidia/Rozella clade. PMID:26715630

  16. CXCR4 gene transfer prevents pressure overload induced heart failure

    PubMed Central

    LaRocca, Thomas J.; Jeong, Dongtak; Kohlbrenner, Erik; Lee, Ahyoung; Chen, JiQiu; Hajjar, Roger J.; Tarzami, Sima T.

    2012-01-01

    Stem cell and gene therapies are being pursued as strategies for repairing damaged cardiac tissue following myocardial infarction in an attempt to prevent heart failure. The chemokine receptor-4 (CXCR4) and its ligand, CXCL12, play a critical role in stem cell recruitment post-acute myocardial infarction. Whereas progenitor cell migration via the CXCL12/CXCR4 axis is well characterized, little is known about the molecular mechanisms of CXCR4 mediated modulation of cardiac hypertrophy and failure. We used gene therapy to test the effects of CXCR4 gene delivery on adverse ventricular remodeling due to pressure overload. We assessed the effect of cardiac overexpression of CXCR4 during trans-aortic constriction (TAC) using a cardiotropic adeno-associated viral vector (AAV9) carrying the CXCR4 gene. Cardiac overexpression of CXCR4 in mice with pressure overload prevented ventricular remodeling, preserved capillary density and maintained function as determined by echocardiography and in vivo hemodynamics. In isolated adult rat cardiac myocytes, CXCL12 treatment prevented isoproterenol induced hypertrophy and interrupted the calcineurin/NFAT pathway. Finally, a complex involving the L-type calcium channel, β2-adenoreceptor, and CXCR4 (Cav1.2/β2AR/CXCR4) was identified in healthy cardiac myocytes and was shown to dissociate as a consequence of heart failure. CXCR4 administered to the heart via gene transfer prevents pressure overload induced heart failure. The identification of CXCR4 participation in a Cav1.2-β2AR regulatory complex provides further insight into the mechanism by which CXCR4 modulates calcium homeostasis and chronic pressure overload responses in the cardiac myocyte. Together these results suggest AAV9.CXCR4 gene therapy is a potential therapeutic approach for congestive heart failure. PMID:22668785

  17. Resistance Gene Transfer during Treatments for Experimental Avian Colibacillosis

    PubMed Central

    Dheilly, Alexandra; Le Devendec, Laëtitia; Mourand, Gwenaëlle; Bouder, Axelle; Jouy, Eric

    2012-01-01

    An experiment was conducted in animal facilities to compare the impacts of four avian colibacillosis treatments—oxytetracycline (OTC), trimethoprim-sulfadimethoxine (SXT), amoxicillin (AMX), or enrofloxacin (ENR)—on the susceptibility of Escherichia coli in broiler intestinal tracts. Birds were first orally inoculated with rifampin-resistant E. coli strains bearing plasmid genes conferring resistance to fluoroquinolones (qnr), cephalosporins (blaCTX-M or blaFOX), trimethoprim-sulfonamides, aminoglycosides, or tetracyclines. Feces samples were collected before, during, and after antimicrobial treatments. The susceptibilities of E. coli strains were studied, and resistance gene transfer was analyzed. An increase in the tetracycline-resistant E. coli population was observed only in OTC-treated birds, whereas multiresistant E. coli was detected in the dominant E. coli populations of SXT-, AMX-, or ENR-treated birds. Most multiresistant E. coli strains were susceptible to rifampin and exhibited various pulsed-field gel electrophoresis profiles, suggesting the transfer of one of the multiresistance plasmids from the inoculated strains to other E. coli strains in the intestinal tract. In conclusion, this study clearly illustrates how, in E. coli, “old” antimicrobials may coselect antimicrobial resistance to recent and critical molecules. PMID:21986830

  18. Lateral gene transfers have polished animal genomes: lessons from nematodes

    PubMed Central

    Danchin, Etienne G. J.; Rosso, Marie-Noëlle

    2012-01-01

    It is now accepted that lateral gene transfers (LGT), have significantly contributed to the composition of bacterial genomes. The amplitude of the phenomenon is considered so high in prokaryotes that it challenges the traditional view of a binary hierarchical tree of life to correctly represent the evolutionary history of species. Given the plethora of transfers between prokaryotes, it is currently impossible to infer the last common ancestral gene set for any extant species. For this ensemble of reasons, it has been proposed that the Darwinian binary tree of life may be inappropriate to correctly reflect the actual relations between species, at least in prokaryotes. In contrast, the contribution of LGT to the composition of animal genomes is less documented. In the light of recent analyses that reported series of LGT events in nematodes, we discuss the importance of this phenomenon in the evolutionary history and in the current composition of an animal genome. Far from being neutral, it appears that besides having contributed to nematode genome contents, LGT have favored the emergence of important traits such as plant-parasitism. PMID:22919619

  19. Synthetic Fatty Acids Prevent Plasmid-Mediated Horizontal Gene Transfer

    PubMed Central

    Getino, María; Sanabria-Ríos, David J.; Fernández-López, Raúl; Campos-Gómez, Javier; Sánchez-López, José M.; Fernández, Antonio; Carballeira, Néstor M.

    2015-01-01

    ABSTRACT Bacterial conjugation constitutes a major horizontal gene transfer mechanism for the dissemination of antibiotic resistance genes among human pathogens. Antibiotic resistance spread could be halted or diminished by molecules that interfere with the conjugation process. In this work, synthetic 2-alkynoic fatty acids were identified as a novel class of conjugation inhibitors. Their chemical properties were investigated by using the prototype 2-hexadecynoic acid and its derivatives. Essential features of effective inhibitors were the carboxylic group, an optimal long aliphatic chain of 16 carbon atoms, and one unsaturation. Chemical modification of these groups led to inactive or less-active derivatives. Conjugation inhibitors were found to act on the donor cell, affecting a wide number of pathogenic bacterial hosts, including Escherichia, Salmonella, Pseudomonas, and Acinetobacter spp. Conjugation inhibitors were active in inhibiting transfer of IncF, IncW, and IncH plasmids, moderately active against IncI, IncL/M, and IncX plasmids, and inactive against IncP and IncN plasmids. Importantly, the use of 2-hexadecynoic acid avoided the spread of a derepressed IncF plasmid into a recipient population, demonstrating the feasibility of abolishing the dissemination of antimicrobial resistances by blocking bacterial conjugation. PMID:26330514

  20. Interleukin-10 Modulates Antigen Presentation by Dendritic Cells through Regulation of NLRP3 Inflammasome Assembly during Chlamydia Infection

    PubMed Central

    Omosun, Yusuf; McKeithen, Danielle; Ryans, Khamia; Kibakaya, Caroline; Blas-Machado, Uriel; Li, Duo; Singh, Rajesh; Inoue, Koichi; Xiong, Zhi-Gang; Eko, Francis; Black, Carolyn; Igietseme, Joseph

    2015-01-01

    Interleukin-10 (IL-10) has been implicated in susceptibility to genital chlamydial infection and the development of tubal pathologies. IL-10 limitation also resulted in the rapid elicitation of immune responses against Chlamydia, and decreased levels of IL-10 correlated with protective anti-Chlamydia immunity. To investigate the molecular basis for these effects, we compared the reproductive pathologies and fertility rates in Chlamydia-infected wild-type (WT) and IL-10-knockout (IL-10−/−) mice; we also analyzed the expression of the Toll-like receptor (TLR)/interleukin-1 receptor (IL-1R) superfamily, IL-1β production, NLRP3 inflammasome assembly and activation, and the immunostimulatory capacity and apoptotic predilection of Chlamydia-exposed dendritic cells (DCs) from WT and IL-10−/− mice. Our results revealed that, in addition to the rapid clearance of infection, genitally infected IL-10−/− mice were protected from tubal pathologies and infertility, whereas WT (IL-10+/+) mice were not. Chlamydia-pulsed IL-10−/− DCs expressed larger numbers of TLR4/IL-1R molecules and had enhanced IL-1β production. In addition, NLRP3 inflammasome assembly was suppressed in IL-10−/− DCs through the inhibition of the P2X purinoceptor 7 (P2X7) receptor (P2X7R), an ATP-gated ion channel, and a decrease in intracellular Ca2+ levels, which inhibited DC apoptosis. Thus, the potent immunostimulatory capacity of IL-10-deficient DCs is due, at least in part, to the suppression of the intracellular inflammasome assembly, which prevents DC apoptosis, allowing efficient antigen presentation. The results indicate that IL-10 deficiency enables efficient antigen presentation by DCs for rapid and enhanced immune activation against Chlamydia, which results in rapid microbial clearance, which prevents tubal pathologies during infection. Our finding has important implications for the induction of protective immunity against Chlamydia and other infectious and noninfectious

  1. Local immunotherapy via delivery of interleukin-10 and transforming growth factor β antagonist for treatment of chronic kidney disease.

    PubMed

    Rodell, Christopher B; Rai, Reena; Faubel, Sarah; Burdick, Jason A; Soranno, Danielle E

    2015-05-28

    Obstructive nephropathy is the leading cause of kidney disease in children. The tissue injury resulting from initial dilation precipitates a deleterious cascade of macrophage infiltration, apoptosis, and fibrosis to produce a resultant dysfunctional tissue. We propose to abate this tissue remodeling process through immunotherapy administered via the local and sustained delivery of interleukin-10 (IL-10; anti-inflammatory) and anti-transforming growth factor β (anti-TGFβ; anti-fibrotic). Shear-thinning, injectable hyaluronic acid (HA) hydrogels were formed through supramolecular guest-host interactions and used to contain IL-10, anti-TGFβ, or both molecules together. Degradation assays demonstrated that diffusive molecule release was associated with concurrent hydrogel erosion and was sustained for up to 3weeks in vitro. Erosion was likewise monitored in vivo by non-invasive optical imaging, where gel localization to the affected tissue was observed with near complete clearance by day 18. Hydrogels were applied to a murine model of chronic kidney disease, with subcapsular hydrogel injections acting as a delivery depot. Quantitative histological analysis (days 7, 21, and 35) was used to evaluate treatment efficacy. Notably, results demonstrated reduced macrophage infiltration beyond day 7 in treatment groups and reduced apoptosis at day 21, relative to untreated unilateral ureteral obstruction disease model. Fibrosis was reduced at the 35day timepoint in groups treated with IL-10 or anti-TGFβ alone, but not with the combination therapy. Rather, dual delivery of IL-10 and anti-TGFβ resulted in a paradoxical hastening of fibrosis, warranting further investigation. Localized immunotherapy is a novel approach to treat kidney disease and shows promise as a translatable therapy. PMID:25804871

  2. Association between interleukin 1β and interleukin 10 concentrations: a cross-sectional study in young adolescents in Taiwan

    PubMed Central

    2013-01-01

    Background In adults, low circulating interleukin 10 (IL10) has been associated with obesity and type 2 diabetes. However, studies investigating IL10 in overweight and obese children have yielded conflicting results. The aim of this study was to investigate factors associated with serum IL10 concentration in young Chinese adolescents. Methods Young adolescents (n=325) ages 13.33±1.10 years were recruited into the cross-sectional study from 2010 to 2011. Parameters of obesity, individual components of MetS, iron status and serum IL10 were evaluated. Results Compared with their normal weight counterparts, overweight adolescents had lower serum IL10 but higher TNFα, nitric oxide (NO) and IL1β concentrations (all p<0.05). Obese adolescents had increased IL1β but decreased hepcidin concentration compared with normal weight (p<0.01 and p<0.05; respectively). A strong inverse relationship (p<0.0001) was found between IL10 and pro-inflammatory cytokines (TNFα and IL1β). Multivariate linear regression analysis showed serum IL1β was significantly correlated with IL10 (β=−0.156, p<0.0001). When overweight and obese adolescents were assessed separately from normal weight, only IL1β was inversely associated with serum IL10 (β=−0.231, p=0.0009). The association between IL10 and IL1β was weaker in adolescents with normal weight (β=−0.157, p=0.0002), after adjusting for gender, TNFα, IFNγ and NO. Conclusions Our study confirmed that low IL10 concentration is associated with overweight and obesity in young adolescents. We also demonstrated for the first time that pro-inflammatory cytokine IL1β is independently associated with IL10. A decline in IL10 concentration in overweight and obese adolescents may further contribute to the IL1β-mediated inflammatory environment associated with obesity. PMID:23941335

  3. Transient activation of mucosal effector immune responses by resident intestinal bacteria in normal hosts is regulated by interleukin-10 signalling.

    PubMed

    Wu, Cong; Sartor, R Balfour; Huang, Kehe; Tonkonogy, Susan L

    2016-07-01

    Interleukin-10 (IL-10) is a key regulator of mucosal homeostasis. In the current study we investigated the early events after monoassociating germ-free (GF) wild-type (WT) mice with an Escherichia coli strain that we isolated previously from the caecal contents of a normal mouse housed under specific pathogen-free conditions. Our results show that interferon-γ (IFN-γ) secreted by mesenteric lymph node (MLN) cells from both IL-10 deficient mice and WT mice, stimulated ex vivo with E. coli lysate, was dramatically higher at day 4 after monoassociation compared with IFN-γ secreted by cells from GF mice without E. coli colonization. Production of IFN-γ rapidly and progressively declined after colonization of WT but not IL-10-deficient mice. The E. coli lysate-stimulated WT MLN cells also produced IL-10 that peaked at day 4 and subsequently declined, but not as precipitously as IFN-γ. WT cells that express CD4, CD8 and NKp46 produced IFN-γ; WT CD4-positive cells and B cells produced IL-10. Recombinant IL-10 added to E. coli-stimulated MLN cell cultures inhibited IFN-γ secretion in a dose-dependent fashion. MLN cells from WT mice treated in vivo with neutralizing anti-IL-10 receptor antibody produced more IFN-γ compared with MLN cells from isotype control antibody-treated mice. These findings show that a resident E. coli that induces chronic colitis in monoassociated IL-10-deficient mice rapidly but transiently activates the effector immune system in normal hosts, in parallel with induction of protective IL-10 produced by B cells and CD4(+) cells that subsequently suppresses this response to mediate mucosal homeostasis. PMID:27147411

  4. Interleukin-10 modulates antigen presentation by dendritic cells through regulation of NLRP3 inflammasome assembly during Chlamydia infection.

    PubMed

    Omosun, Yusuf; McKeithen, Danielle; Ryans, Khamia; Kibakaya, Caroline; Blas-Machado, Uriel; Li, Duo; Singh, Rajesh; Inoue, Koichi; Xiong, Zhi-Gang; Eko, Francis; Black, Carolyn; Igietseme, Joseph; He, Qing

    2015-12-01

    Interleukin-10 (IL-10) has been implicated in susceptibility to genital chlamydial infection and the development of tubal pathologies. IL-10 limitation also resulted in the rapid elicitation of immune responses against Chlamydia, and decreased levels of IL-10 correlated with protective anti-Chlamydia immunity. To investigate the molecular basis for these effects, we compared the reproductive pathologies and fertility rates in Chlamydia-infected wild-type (WT) and IL-10-knockout (IL-10(-/-)) mice; we also analyzed the expression of the Toll-like receptor (TLR)/interleukin-1 receptor (IL-1R) superfamily, IL-1β production, NLRP3 inflammasome assembly and activation, and the immunostimulatory capacity and apoptotic predilection of Chlamydia-exposed dendritic cells (DCs) from WT and IL-10(-/-) mice. Our results revealed that, in addition to the rapid clearance of infection, genitally infected IL-10(-/-) mice were protected from tubal pathologies and infertility, whereas WT (IL-10(+/+)) mice were not. Chlamydia-pulsed IL-10(-/-) DCs expressed larger numbers of TLR4/IL-1R molecules and had enhanced IL-1β production. In addition, NLRP3 inflammasome assembly was suppressed in IL-10(-/-) DCs through the inhibition of the P2X purinoceptor 7 (P2X7) receptor (P2X7R), an ATP-gated ion channel, and a decrease in intracellular Ca(2+) levels, which inhibited DC apoptosis. Thus, the potent immunostimulatory capacity of IL-10-deficient DCs is due, at least in part, to the suppression of the intracellular inflammasome assembly, which prevents DC apoptosis, allowing efficient antigen presentation. The results indicate that IL-10 deficiency enables efficient antigen presentation by DCs for rapid and enhanced immune activation against Chlamydia, which results in rapid microbial clearance, which prevents tubal pathologies during infection. Our finding has important implications for the induction of protective immunity against Chlamydia and other infectious and noninfectious diseases by

  5. Induction of monocyte chemoattractant protein-1 and interleukin-10 by TGFbeta1 in melanoma enhances tumor infiltration and immunosuppression.

    PubMed

    Díaz-Valdés, Nancy; Basagoiti, María; Dotor, Javier; Aranda, Fernando; Monreal, Iñaki; Riezu-Boj, José Ignacio; Borrás-Cuesta, Francisco; Sarobe, Pablo; Feijoó, Esperanza

    2011-02-01

    Melanoma progression is associated with the expression of different growth factors, cytokines, and chemokines. Because TGFβ1 is a pleiotropic cytokine involved not only in physiologic processes but also in cancer development, we analyzed in A375 human melanoma cells, the effect of TGFβ1 on monocyte chemoattractant protein-1 (MCP-1) and interleukin-10 (IL-10) expression, two known factors responsible for melanoma progression. TGFβ1 increased the expression of MCP-1 and IL-10 in A375 cells, an effect mediated by the cross-talk between Smad, PI3K (phosphoinositide 3-kinase)/AKT, and BRAF-MAPK (mitogen activated protein kinase) signaling pathways. Supernatants from TGFβ1-treated A375 cells enhanced MCP-1-dependent migration of monocytes, which, in turn, expressed high levels of TGF,β1, bFGF, and VEGF mRNA. Moreover, these supernatants also inhibited functional properties of dendritic cells through IL-10-dependent mechanisms. When using in vitro, the TGFβ1-blocking peptide P144, TGFβ1-dependent Smad3 phosphorylation, and expression of MCP-1 and IL-10 were inhibited. In vivo, treatment of A375 tumor-bearing athymic mice with P144 significantly reduced tumor growth, associated with a lower macrophage infiltrate and decreased intratumor MCP-1 and VEGF levels, as well as angiogenesis. Finally, in C57BL/6 mice with B16-OVA melanoma tumors, when administered with immunotherapy, P144 decreased tumor growth and intratumor IL-10 levels, linked to enhanced activation of dendritic cells and natural killer cells, as well as anti-OVA T-cell responses. These results show new effects of TGFβ1 on melanoma cells, which promote tumor progression and immunosuppression, strongly reinforcing the relevance of this cytokine as a molecular target in melanoma. PMID:21159663

  6. Viral Infection of the Central Nervous System Exacerbates Interleukin-10 Receptor Deficiency-Mediated Colitis in SJL Mice.

    PubMed

    Uhde, Ann-Kathrin; Herder, Vanessa; Akram Khan, Muhammad; Ciurkiewicz, Malgorzata; Schaudien, Dirk; Teich, René; Floess, Stefan; Baumgärtner, Wolfgang; Huehn, Jochen; Beineke, Andreas

    2016-01-01

    Theiler´s murine encephalomyelitis virus (TMEV)-infection is a widely used animal model for studying demyelinating disorders, including multiple sclerosis (MS). The immunosuppressive cytokine Interleukin (IL)-10 counteracts hyperactive immune responses and critically controls immune homeostasis in infectious and autoimmune disorders. In order to investigate the effect of signaling via Interleukin-10 receptor (IL-10R) in infectious neurological diseases, TMEV-infected SJL mice were treated with IL-10R blocking antibody (Ab) in the acute and chronic phase of the disease. The findings demonstrate that (i) Ab-mediated IL-10 neutralization leads to progressive colitis with a reduction in Foxp3+ regulatory T cells and increased numbers of CD8+CD44+ memory T cells as well as activated CD4+CD69+ and CD8+CD69+ T cells in uninfected mice. (ii) Concurrent acute TMEV-infection worsened enteric disease-mediated by IL-10R neutralization. Virus-triggered effects were associated with an enhanced activation of CD4+ T helper cells and CD8+ cytotoxic T lymphocytes and augmented cytokine expression. By contrast, (iii) IL-10R neutralization during chronic TMEV-infection was not associated with enhanced peripheral immunopathology but an increased CD3+ T cell influx in the spinal cord. IL-10R neutralization causes a breakdown in peripheral immune tolerance in genetically predisposed mice, which leads to immune-mediated colitis, resembling inflammatory bowel disease. Hyperactive immune state following IL-10R blockade is enhanced by central nervous system-restricted viral infection in a disease phase-dependent manner. PMID:27611574

  7. Amoebozoa possess lineage-specific globin gene repertoires gained by individual horizontal gene transfers.

    PubMed

    Dröge, Jasmin; Buczek, Dorota; Suzuki, Yutaka; Makałowski, Wojciech

    2014-01-01

    The Amoebozoa represent a clade of unicellular amoeboid organisms that display a wide variety of lifestyles, including free-living and parasitic species. For example, the social amoeba Dictyostelium discoideum has the ability to aggregate into a multicellular fruiting body upon starvation, while the pathogenic amoeba Entamoeba histolytica is a parasite of humans. Globins are small heme proteins that are present in almost all extant organisms. Although several genomes of amoebozoan species have been sequenced, little is known about the phyletic distribution of globin genes within this phylum. Only two flavohemoglobins (FHbs) of D. discoideum have been reported and characterized previously while the genomes of Entamoeba species are apparently devoid of globin genes. We investigated eleven amoebozoan species for the presence of globin genes by genomic and phylogenetic in silico analyses. Additional FHb genes were identified in the genomes of four social amoebas and the true slime mold Physarum polycephalum. Moreover, a single-domain globin (SDFgb) of Hartmannella vermiformis, as well as two truncated hemoglobins (trHbs) of Acanthamoeba castellanii were identified. Phylogenetic evidence suggests that these globin genes were independently acquired via horizontal gene transfer from some ancestral bacteria. Furthermore, the phylogenetic tree of amoebozoan FHbs indicates that they do not share a common ancestry and that a transfer of FHbs from bacteria to amoeba occurred multiple times. PMID:25013378

  8. Genome-wide experimental determination of barriers to horizontal gene transfer.

    PubMed

    Sorek, Rotem; Zhu, Yiwen; Creevey, Christopher J; Francino, M Pilar; Bork, Peer; Rubin, Edward M

    2007-11-30

    Horizontal gene transfer, in which genetic material is transferred from the genome of one organism to that of another, has been investigated in microbial species mainly through computational sequence analyses. To address the lack of experimental data, we studied the attempted movement of 246,045 genes from 79 prokaryotic genomes into Escherichia coli and identified genes that consistently fail to transfer. We studied the mechanisms underlying transfer inhibition by placing coding regions from different species under the control of inducible promoters. Our data suggest that toxicity to the host inhibited transfer regardless of the species of origin and that increased gene dosage and associated increased expression may be a predominant cause for transfer failure. Although these experimental studies examined transfer solely into E. coli, a computational analysis of gene-transfer rates across available bacterial and archaeal genomes supports that the barriers observed in our study are general across the tree of life. PMID:17947550

  9. Genome-wide experimental determination of barriers to horizontal gene transfer

    SciTech Connect

    Rubin, Edward; Sorek, Rotem; Zhu, Yiwen; Creevey, Christopher J.; Francino, M. Pilar; Bork, Peer; Rubin, Edward M.

    2007-09-24

    Horizontal gene transfer, in which genetic material is transferred from the genome of one organism to another, has been investigated in microbial species mainly through computational sequence analyses. To address the lack of experimental data, we studied the attempted movement of 246,045 genes from 79 prokaryotic genomes into E. coli and identified genes that consistently fail to transfer. We studied the mechanisms underlying transfer inhibition by placing coding regions from different species under the control of inducible promoters. Their toxicity to the host inhibited transfer regardless of the species of origin and our data suggest that increased gene dosage and associated increased expression is a predominant cause for transfer failure. While these experimental studies examined transfer solely into E. coli, a computational analysis of gene transfer rates across available bacterial and archaeal genomes indicates that the barriers observed in our study are general across the tree of life.

  10. Repeated, recent and diverse transfers of a mitochondrial gene to the nucleus in flowering plants.

    PubMed

    Adams, K L; Daley, D O; Qiu, Y L; Whelan, J; Palmer, J D

    2000-11-16

    A central component of the endosymbiotic theory for the bacterial origin of the mitochondrion is that many of its genes were transferred to the nucleus. Most of this transfer occurred early in mitochondrial evolution; functional transfer of mitochondrial genes has ceased in animals. Although mitochondrial gene transfer continues to occur in plants, no comprehensive study of the frequency and timing of transfers during plant evolution has been conducted. Here we report frequent loss (26 times) and transfer to the nucleus of the mitochondrial gene rps10 among 277 diverse angiosperms. Characterization of nuclear rps10 genes from 16 out of 26 loss lineages implies that many independent, RNA-mediated rps10 transfers occurred during recent angiosperm evolution; each of the genes may represent a separate functional gene transfer. Thus, rps10 has been transferred to the nucleus at a surprisingly high rate during angiosperm evolution. The structures of several nuclear rps10 genes reveal diverse mechanisms by which transferred genes become activated, including parasitism of pre-existing nuclear genes for mitochondrial or cytoplasmic proteins, and activation without gain of a mitochondrial targeting sequence. PMID:11099041

  11. Adaptive Horizontal Gene Transfers between Multiple Cheese-Associated Fungi

    PubMed Central

    Ropars, Jeanne; Rodríguez de la Vega, Ricardo C.; López-Villavicencio, Manuela; Gouzy, Jérôme; Sallet, Erika; Dumas, Émilie; Lacoste, Sandrine; Debuchy, Robert; Dupont, Joëlle; Branca, Antoine; Giraud, Tatiana

    2015-01-01

    Summary Domestication is an excellent model for studies of adaptation because it involves recent and strong selection on a few, identified traits [1–5]. Few studies have focused on the domestication of fungi, with notable exceptions [6–11], despite their importance to bioindustry [12] and to a general understanding of adaptation in eukaryotes [5]. Penicillium fungi are ubiquitous molds among which two distantly related species have been independently selected for cheese making—P. roqueforti for blue cheeses like Roquefort and P. camemberti for soft cheeses like Camembert. The selected traits include morphology, aromatic profile, lipolytic and proteolytic activities, and ability to grow at low temperatures, in a matrix containing bacterial and fungal competitors [13–15]. By comparing the genomes of ten Penicillium species, we show that adaptation to cheese was associated with multiple recent horizontal transfers of large genomic regions carrying crucial metabolic genes. We identified seven horizontally transferred regions (HTRs) spanning more than 10 kb each, flanked by specific transposable elements, and displaying nearly 100% identity between distant Penicillium species. Two HTRs carried genes with functions involved in the utilization of cheese nutrients or competition and were found nearly identical in multiple strains and species of cheese-associated Penicillium fungi, indicating recent selective sweeps; they were experimentally associated with faster growth and greater competitiveness on cheese and contained genes highly expressed in the early stage of cheese maturation. These findings have industrial and food safety implications and improve our understanding of the processes of adaptation to rapid environmental changes. PMID:26412136

  12. Endogenous mouse interleukin-10 is up-regulated by exogenously administered recombinant human interleukin-10, but does not contribute to the efficacy of the human protein in mouse models of endotoxemia.

    PubMed

    Smith, S R; Terminelli, C; Denhardt, G; Manfra, D; Davies, L; Narula, S

    1999-02-01

    In murine models of experimental endotoxemia, inflammatory cytokines as well as antiinflammatory interleukin-10 (IL-10) appear in the circulation after the injection of lipopolysaccharide (LPS). There is considerable experimental evidence to suggest that the major function of endogenously produced IL-10 is to down-regulate inflammatory cytokine production. Indeed, the protective effects of exogenously administered IL-10 against murine endotoxin lethality have been shown to correlate with its ability to inhibit the LPS-induced production of tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma). While mouse IL-10 (mIL-10) has been used in the majority of studies in murine endotoxemia, we have found the human homolog to be equally effective in suppressing inflammatory cytokine production and in protecting mice from endotoxin lethality. However, we have recently observed that the LPS-induced endogenous IL-10 response is enhanced when mice are treated with recombinant human IL-10 (rhuIL-10). The upregulation of endogenous IL-10 by exogenously administered rhuIL-10 is particularly evident in mice that are primed with Corynebacterium partum (Proprionibacterium acnes). In the present study, we have examined the potential contributions of the increased circulating levels of mouse IL-10 to the inhibitory effects seen with rhuIL-10 on inflammatory cytokine production and endotoxin lethality. We show that pretreatment with a neutralizing anti-mouse IL-10 monoclonal antibody (mAb) has no effect on the ability of rhuIL-10 to suppress an LPS-induced inflammatory cytokine response in these mice. In contrast, the suppressive effects of the human protein on inflammatory cytokine responses are blocked completely by pretreating the animals with an anti-huIL-10 mAb. These data show that despite the up-regulated endogenous IL-10 response, it is the exogenously administered rhuIL-10 that is directly responsible for the suppressed inflammatory cytokine responses that

  13. Extensive Intra-Kingdom Horizontal Gene Transfer Converging on a Fungal Fructose Transporter Gene

    PubMed Central

    Coelho, Marco A.; Gonçalves, Carla; Sampaio, José Paulo; Gonçalves, Paula

    2013-01-01

    Comparative genomics revealed in the last decade a scenario of rampant horizontal gene transfer (HGT) among prokaryotes, but for fungi a clearly dominant pattern of vertical inheritance still stands, punctuated however by an increasing number of exceptions. In the present work, we studied the phylogenetic distribution and pattern of inheritance of a fungal gene encoding a fructose transporter (FSY1) with unique substrate selectivity. 109 FSY1 homologues were identified in two sub-phyla of the Ascomycota, in a survey that included 241 available fungal genomes. At least 10 independent inter-species instances of horizontal gene transfer (HGT) involving FSY1 were identified, supported by strong phylogenetic evidence and synteny analyses. The acquisition of FSY1 through HGT was sometimes suggestive of xenolog gene displacement, but several cases of pseudoparalogy were also uncovered. Moreover, evidence was found for successive HGT events, possibly including those responsible for transmission of the gene among yeast lineages. These occurrences do not seem to be driven by functional diversification of the Fsy1 proteins because Fsy1 homologues from widely distant lineages, including at least one acquired by HGT, appear to have similar biochemical properties. In summary, retracing the evolutionary path of the FSY1 gene brought to light an unparalleled number of independent HGT events involving a single fungal gene. We propose that the turbulent evolutionary history of the gene may be linked to the unique biochemical properties of the encoded transporter, whose predictable effect on fitness may be highly variable. In general, our results support the most recent views suggesting that inter-species HGT may have contributed much more substantially to shape fungal genomes than heretofore assumed. PMID:23818872

  14. Applying horizontal gene transfer phenomena to enhance non-viral gene therapy

    PubMed Central

    Elmer, Jacob J.; Christensen, Matthew D.; Rege, Kaushal

    2014-01-01

    Horizontal gene transfer (HGT) is widespread amongst prokaryotes, but eukaryotes tend to be far less promiscuous with their genetic information. However, several examples of HGT from pathogens into eukaryotic cells have been discovered and mimicked to improve non-viral gene delivery techniques. For example, several viral proteins and DNA sequences have been used to significantly increase cytoplasmic and nuclear gene delivery. Plant genetic engineering is routinely performed with the pathogenic bacterium Agrobacterium tumefaciens and similar pathogens (e.g. Bartonella henselae) may also be able to transform human cells. Intracellular parasites like Trypanosoma cruzi may also provide new insights into overcoming cellular barriers to gene delivery. Finally, intercellular nucleic acid transfer between host cells will also be briefly discussed. This article will review the unique characteristics of several different viruses and microbes and discuss how their traits have been successfully applied to improve non-viral gene delivery techniques. Consequently, pathogenic traits that originally caused diseases may eventually be used to treat many genetic diseases. PMID:23994344

  15. Comparison between Agrobacterium-mediated and direct gene transfer using the gene gun.

    PubMed

    Gao, Caixia; Nielsen, Klaus K

    2013-01-01

    Agrobacterium-mediated transformation and direct gene transfer using the gene gun (microparticle -bombardment) are the two most widely used methods for plant genetic modification. The Agrobacterium method has been successfully practiced in dicots for many years, but only recently have efficient protocols been developed for grasses. Microparticle bombardment has evolved as a method delivering exogenous nucleic acids into plant genome and is a commonly employed technique in plant science. Here these two systems are compared for transformation efficiency, transgene integration, and transgene expression when used to transform tall fescue (Festuca arundinacea Schreb.). The tall fescue transformation protocols lead to the production of large numbers of fertile, independent transgenic lines. PMID:23104329

  16. Multiple Inter-Kingdom Horizontal Gene Transfers in the Evolution of the Phosphoenolpyruvate Carboxylase Gene Family

    PubMed Central

    Wang, Wen; Su, Bing

    2012-01-01

    Pepcase is a gene encoding phosphoenolpyruvate carboxylase that exists in bacteria, archaea and plants,playing an important role in plant metabolism and development. Most plants have two or more pepcase genes belonging to two gene sub-families, while only one gene exists in other organisms. Previous research categorized one plant pepcase gene as plant-type pepcase (PTPC) while the other as bacteria-type pepcase (BTPC) because of its similarity with the pepcase gene found in bacteria. Phylogenetic reconstruction showed that PTPC is the ancestral lineage of plant pepcase, and that all bacteria, protistpepcase and BTPC in plants are derived from a lineage of pepcase closely related with PTPC in algae. However, their phylogeny contradicts the species tree and traditional chronology of organism evolution. Because the diversification of bacteria occurred much earlier than the origin of plants, presumably all bacterialpepcase derived from the ancestral PTPC of algal plants after divergingfrom the ancestor of vascular plant PTPC. To solve this contradiction, we reconstructed the phylogeny of pepcase gene family. Our result showed that both PTPC and BTPC are derived from an ancestral lineage of gamma-proteobacteriapepcases, possibly via an ancient inter-kingdom horizontal gene transfer (HGT) from bacteria to the eukaryotic common ancestor of plants, protists and cellular slime mold. Our phylogenetic analysis also found 48other pepcase genes originated from inter-kingdom HGTs. These results imply that inter-kingdom HGTs played important roles in the evolution of the pepcase gene family and furthermore that HGTsare a more frequent evolutionary event than previouslythought. PMID:23251445

  17. Multiple inter-kingdom horizontal gene transfers in the evolution of the phosphoenolpyruvate carboxylase gene family.

    PubMed

    Peng, Yingmei; Cai, Jing; Wang, Wen; Su, Bing

    2012-01-01

    Pepcase is a gene encoding phosphoenolpyruvate carboxylase that exists in bacteria, archaea and plants,playing an important role in plant metabolism and development. Most plants have two or more pepcase genes belonging to two gene sub-families, while only one gene exists in other organisms. Previous research categorized one plant pepcase gene as plant-type pepcase (PTPC) while the other as bacteria-type pepcase (BTPC) because of its similarity with the pepcase gene found in bacteria. Phylogenetic reconstruction showed that PTPC is the ancestral lineage of plant pepcase, and that all bacteria, protistpepcase and BTPC in plants are derived from a lineage of pepcase closely related with PTPC in algae. However, their phylogeny contradicts the species tree and traditional chronology of organism evolution. Because the diversification of bacteria occurred much earlier than the origin of plants, presumably all bacterialpepcase derived from the ancestral PTPC of algal plants after divergingfrom the ancestor of vascular plant PTPC. To solve this contradiction, we reconstructed the phylogeny of pepcase gene family. Our result showed that both PTPC and BTPC are derived from an ancestral lineage of gamma-proteobacteriapepcases, possibly via an ancient inter-kingdom horizontal gene transfer (HGT) from bacteria to the eukaryotic common ancestor of plants, protists and cellular slime mold. Our phylogenetic analysis also found 48other pepcase genes originated from inter-kingdom HGTs. These results imply that inter-kingdom HGTs played important roles in the evolution of the pepcase gene family and furthermore that HGTsare a more frequent evolutionary event than previouslythought. PMID:23251445

  18. Improved gene transfer with histidine-functionalized mesoporous silica nanoparticles.

    PubMed

    Brevet, David; Hocine, Ouahiba; Delalande, Anthony; Raehm, Laurence; Charnay, Clarence; Midoux, Patrick; Durand, Jean-Olivier; Pichon, Chantal

    2014-08-25

    Mesoporous silica nanoparticles (MSN) were functionalized with aminopropyltriethoxysilane (MSN-NH2) then L-histidine (MSN-His) for pDNA delivery in cells and in vivo. The complexation of pDNA with MSN-NH2 and MSN-His was first studied with gel shift assay. pDNA complexed with MSN-His was better protected from DNase degradation than with MSN-NH2. An improvement of the transfection efficiency in cells was observed with MSN-His/pDNA compared to MSN-NH2/pDNA, which could be explained by a better internalization of MSN-His. The improvement of the transfection efficiency with MSN-His was also observed for gene transfer in Achilles tendons in vivo. PMID:24853464

  19. Statistical Mechanics of Horizontal Gene Transfer in Evolutionary Ecology

    NASA Astrophysics Data System (ADS)

    Chia, Nicholas; Goldenfeld, Nigel

    2011-04-01

    The biological world, especially its majority microbial component, is strongly interacting and may be dominated by collective effects. In this review, we provide a brief introduction for statistical physicists of the way in which living cells communicate genetically through transferred genes, as well as the ways in which they can reorganize their genomes in response to environmental pressure. We discuss how genome evolution can be thought of as related to the physical phenomenon of annealing, and describe the sense in which genomes can be said to exhibit an analogue of information entropy. As a direct application of these ideas, we analyze the variation with ocean depth of transposons in marine microbial genomes, predicting trends that are consistent with recent observations using metagenomic surveys.

  20. Detecting rare gene transfer events in bacterial populations.

    PubMed

    Nielsen, Kaare M; Bøhn, Thomas; Townsend, Jeffrey P

    2014-01-01

    Horizontal gene transfer (HGT) enables bacteria to access, share, and recombine genetic variation, resulting in genetic diversity that cannot be obtained through mutational processes alone. In most cases, the observation of evolutionary successful HGT events relies on the outcome of initially rare events that lead to novel functions in the new host, and that exhibit a positive effect on host fitness. Conversely, the large majority of HGT events occurring in bacterial populations will go undetected due to lack of replication success of transformants. Moreover, other HGT events that would be highly beneficial to new hosts can fail to ensue due to lack of physical proximity to the donor organism, lack of a suitable gene transfer mechanism, genetic compatibility, and stochasticity in tempo-spatial occurrence. Experimental attempts to detect HGT events in bacterial populations have typically focused on the transformed cells or their immediate offspring. However, rare HGT events occurring in large and structured populations are unlikely to reach relative population sizes that will allow their immediate identification; the exception being the unusually strong positive selection conferred by antibiotics. Most HGT events are not expected to alter the likelihood of host survival to such an extreme extent, and will confer only minor changes in host fitness. Due to the large population sizes of bacteria and the time scales involved, the process and outcome of HGT are often not amenable to experimental investigation. Population genetic modeling of the growth dynamics of bacteria with differing HGT rates and resulting fitness changes is therefore necessary to guide sampling design and predict realistic time frames for detection of HGT, as it occurs in laboratory or natural settings. Here we review the key population genetic parameters, consider their complexity and highlight knowledge gaps for further research. PMID:24432015

  1. Detecting rare gene transfer events in bacterial populations

    PubMed Central

    Nielsen, Kaare M.; Bøhn, Thomas; Townsend, Jeffrey P.

    2014-01-01

    Horizontal gene transfer (HGT) enables bacteria to access, share, and recombine genetic variation, resulting in genetic diversity that cannot be obtained through mutational processes alone. In most cases, the observation of evolutionary successful HGT events relies on the outcome of initially rare events that lead to novel functions in the new host, and that exhibit a positive effect on host fitness. Conversely, the large majority of HGT events occurring in bacterial populations will go undetected due to lack of replication success of transformants. Moreover, other HGT events that would be highly beneficial to new hosts can fail to ensue due to lack of physical proximity to the donor organism, lack of a suitable gene transfer mechanism, genetic compatibility, and stochasticity in tempo-spatial occurrence. Experimental attempts to detect HGT events in bacterial populations have typically focused on the transformed cells or their immediate offspring. However, rare HGT events occurring in large and structured populations are unlikely to reach relative population sizes that will allow their immediate identification; the exception being the unusually strong positive selection conferred by antibiotics. Most HGT events are not expected to alter the likelihood of host survival to such an extreme extent, and will confer only minor changes in host fitness. Due to the large population sizes of bacteria and the time scales involved, the process and outcome of HGT are often not amenable to experimental investigation. Population genetic modeling of the growth dynamics of bacteria with differing HGT rates and resulting fitness changes is therefore necessary to guide sampling design and predict realistic time frames for detection of HGT, as it occurs in laboratory or natural settings. Here we review the key population genetic parameters, consider their complexity and highlight knowledge gaps for further research. PMID:24432015

  2. Association of interleukin-10 promoter haplotypes with disease susceptibility and IL-10 levels in Mexican patients with systemic lupus erythematosus.

    PubMed

    Palafox-Sánchez, Claudia Azucena; Oregon-Romero, Edith; Salazar-Camarena, Diana Celeste; Valle, Yeminia Maribel; Machado-Contreras, Jesús René; Cruz, Alvaro; Orozco-López, Mariana; Orozco-Barocio, Gerardo; Vázquez-Del Mercado, Mónica; Muñoz-Valle, José Francisco

    2015-11-01

    Systemic lupus erythematosus (SLE) is the prototype autoimmune rheumatic disease. The etiology of this disease is incompletely understood; however, environmental factors and genetic predisposition are involved. Cytokine-mediated immunity plays a crucial role in the pathogenesis of SLE. We investigate the association of interleukin-10 (IL-10) promoter polymorphisms and their haplotypes in SLE patients from the western Mexico. One hundred and twenty-five SLE patients fulfilling the 1997 ACR criteria and 260 unrelated healthy subjects (HS), both Mexican mestizos, were genotyped for IL-10 -1082A>G, -819C>T, and -592C>A polymorphisms. Haplotypes were inferred using the expectation-maximization algorithm, then allele and haplotype distributions were compared between patients and HS, as well as patients with different clinical variables. We identified at -1082, -819, and -592 four predominant haplotypes ACC (43.70 % in patients vs 46.55 % in HS), ATA (21.45 vs 22.97 %), GCC (16.28 vs 14.21 %), and GTA (14.12 vs 14.12 %). The ATC haplotype was more frequent in SLE respect to HS, suggesting a risk effect (3.23 vs 1.05 %; OR 3.55, CI 1.14-11.11; p = 0.0293). SLE patient carriers of -592 CC genotype as well as the dominant model of inheritance showed higher sIL-10 respect to AA genotype, suggesting that -592 C allele is associated with increased production of the cytokine (p < 0.05). The ACC haplotype had higher IL-10 serum levels and higher values of Mexican version of the Systemic Lupus Erythematosus Disease Activity Index compared with the other haplotype carriers; however, no association was found regarding autoantibodies. Our data suggest that the IL-10 promoter haplotypes play an important role in the risk of developing SLE and influence the production of IL-10 in Mexican population. Nevertheless, further studies are required to analyze the expression of mRNA as well as to investigate the interacting epigenetic factors that could help to define the true contribution of

  3. Interleukin-10 neutralizing antibody for detection of intestinal luminal levels and as a dietary additive in Eimeria challenged broiler chicks.

    PubMed

    Arendt, Maria K; Sand, Jordan M; Marcone, Taylor M; Cook, Mark E

    2016-02-01

    Interleukin-10 (IL-10) mRNA levels are increased within intestinal mucosa after Eimeria infection. IL-10 apical receptor presence on enterocytes suggests IL-10 is secreted into the intestinal lumen. Increased IL-10 has been shown to be central to the pathogenesis of numerous intracellular pathogens; we hypothesize luminal secretion of IL-10 enables Eimeria spp. infection in chickens. This study examines intestine luminal IL-10 levels and performance in broilers challenged with Eimeria when fed an anti-IL-10 antibody. Chicks were fed a diet (1 to 21 d) with control or anti-IL-10 antibody (0.34 g egg yolk antibody powder/Kg diet) with a saline or 10× dose of Advent coccidiosis vaccine on d 3. One chick per pen was euthanized on days 2, 4, 7, 10, 13, 16, and 19 post-challenge, bled, and intestines were collected for luminal fluid IL-10 concentrations. Body weight and feed intake were measured on d 21, and oocyst shedding was assessed on d 7 post-challenge. A significant Eimeria × antibody interaction on d 21 body weight (P < 0.05) showed chicks fed control antibody, but not anti-IL-10, had significant reductions in body weight when challenged with Eimeria spp. Oocyst shedding was increased with Eimeria challenge, but dietary antibody had no effect. Plasma carotenoid levels were reduced in Eimeria challenged chicks 4, 7, 10, and 16 days post-challenge compared to unchallenged chicks. Lack of an Eimeria × antibody interaction showed anti-IL-10 was not protective against Eimeria-induced decreases in plasma carotenoids. Eimeria challenge increased intestine luminal IL-10 on days 4 and 7 post-challenge in the cecum and jejunum, respectively, compared to unchallenged. Dietary anti-IL-10 decreased luminal IL-10 in the ileum on day 2 post-challenge when compared to control antibody fed chicks. No interaction between Eimeria challenge and antibody was observed on intestine luminal contents of IL-10, suggesting anti-IL-10 was ineffective at preventing increased Eimeria

  4. Widespread impact of horizontal gene transfer on plant colonization of land

    PubMed Central

    Yue, Jipei; Hu, Xiangyang; Sun, Hang; Yang, Yongping; Huang, Jinling

    2012-01-01

    In complex multicellular eukaryotes such as animals and plants, horizontal gene transfer is commonly considered rare with very limited evolutionary significance. Here we show that horizontal gene transfer is a dynamic process occurring frequently in the early evolution of land plants. Our genome analyses of the moss Physcomitrella patens identified 57 families of nuclear genes that were acquired from prokaryotes, fungi or viruses. Many of these gene families were transferred to the ancestors of green or land plants. Available experimental evidence shows that these anciently acquired genes are involved in some essential or plant-specific activities such as xylem formation, plant defence, nitrogen recycling as well as the biosynthesis of starch, polyamines, hormones and glutathione. These findings suggest that horizontal gene transfer had a critical role in the transition of plants from aquatic to terrestrial environments. On the basis of these findings, we propose a model of horizontal gene transfer mechanism in nonvascular and seedless vascular plants. PMID:23093189

  5. Multiple losses and transfers to the nucleus of two mitochondrial succinate dehydrogenase genes during angiosperm evolution.

    PubMed Central

    Adams, K L; Rosenblueth, M; Qiu, Y L; Palmer, J D

    2001-01-01

    Unlike in animals, the functional transfer of mitochondrial genes to the nucleus is an ongoing process in plants. All but one of the previously reported transfers in angiosperms involve ribosomal protein genes. Here we report frequent transfer of two respiratory genes, sdh3 and sdh4 (encoding subunits 3 and 4 of succinate dehydrogenase), and we also show that these genes are present and expressed in the mitochondria of diverse angiosperms. Southern hybridization surveys reveal that sdh3 and sdh4 have been lost from the mitochondrion about 40 and 19 times, respectively, among the 280 angiosperm genera examined. Transferred, functional copies of sdh3 and sdh4 were characterized from the nucleus in four and three angiosperm families, respectively. The mitochondrial targeting presequences of two sdh3 genes are derived from preexisting genes for anciently transferred mitochondrial proteins. On the basis of the unique presequences of the nuclear genes and the recent mitochondrial gene losses, we infer that each of the seven nuclear sdh3 and sdh4 genes was derived from a separate transfer to the nucleus. These results strengthen the hypothesis that angiosperms are experiencing a recent evolutionary surge of mitochondrial gene transfer to the nucleus and reveal that this surge includes certain respiratory genes in addition to ribosomal protein genes. PMID:11454775

  6. Kidney-specific Sonoporation-mediated Gene Transfer

    PubMed Central

    Ishida, Ryo; Kami, Daisuke; Kusaba, Tetsuro; Kirita, Yuhei; Kishida, Tsunao; Mazda, Osam; Adachi, Takaomi; Gojo, Satoshi

    2016-01-01

    Sonoporation can deliver agents to target local organs by systemic administration, while decreasing the associated risk of adverse effects. Sonoporation has been used for a variety of materials and in a variety of organs. Herein, we demonstrated that local sonoporation to the kidney can offer highly efficient transfer of oligonucleotides, which were systemically administrated to the tubular epithelium with high specificity. Ultrasonic wave irradiation to the kidney collapsed the microbubbles and transiently affected the glomerular filtration barrier and increased glomerular permeability. Oligonucleotides were passed through the barrier all at once and were absorbed throughout the tubular epithelium. Tumor necrosis factor alpha (TNFα), which plays a central role in renal ischemia–reperfusion injury, was targeted using small interfering RNA (siRNA) with renal sonoporation in a murine model. The reduction of TNFα expression after single gene transfer significantly inhibited the expression of kidney injury markers, suggesting that systemic administration of siRNA under temporary and local sonoporation could be applicable in the clinical setting of ischemic acute kidney injury. PMID:26419704

  7. Center for Fetal Monkey Gene Transfer for Heart, Lung, and Blood Diseases: An NHLBI Resource for the Gene Therapy Community

    PubMed Central

    Skarlatos, Sonia I.

    2012-01-01

    Abstract The goals of the National Heart, Lung, and Blood Institute (NHLBI) Center for Fetal Monkey Gene Transfer for Heart, Lung, and Blood Diseases are to conduct gene transfer studies in monkeys to evaluate safety and efficiency; and to provide NHLBI-supported investigators with expertise, resources, and services to actively pursue gene transfer approaches in monkeys in their research programs. NHLBI-supported projects span investigators throughout the United States and have addressed novel approaches to gene delivery; “proof-of-principle”; assessed whether findings in small-animal models could be demonstrated in a primate species; or were conducted to enable new grant or IND submissions. The Center for Fetal Monkey Gene Transfer for Heart, Lung, and Blood Diseases successfully aids the gene therapy community in addressing regulatory barriers, and serves as an effective vehicle for advancing the field. PMID:22974119

  8. The tryptophanase gene cluster of Haemophilus influenzae type b: evidence for horizontal gene transfer.

    PubMed

    Martin, K; Morlin, G; Smith, A; Nordyke, A; Eisenstark, A; Golomb, M

    1998-01-01

    Among strains of Haemophilus influenzae, the ability to catabolize tryptophan (as detected by indole production) varies and is correlated with pathogenicity. Tryptophan catabolism is widespread (70 to 75%) among harmless respiratory isolates but is nearly universal (94 to 100%) among strains causing serious disease, including meningitis. As a first step in investigating the relationship between tryptophan catabolism and virulence, we have identified genes in pathogenic H. influenzae which are homologous to the tryptophanase (tna) operon of Escherichia coli. The tna genes are located on a 3.1-kb fragment between nlpD and mutS in the H. influenzae type b (Eagan) genome, are flanked by 43-bp direct repeats of an uptake signal sequence downstream from nlpD, and appear to have been inserted as a mobile unit within this sequence. The organization of this insertion is reminiscent of pathogenicity islands. The tna cluster is found at the same map location in all indole-positive strains of H. influenzae surveyed and is absent from reference type d and e genomes. In contrast to H. influenzae, most other Haemophilus species lack tna genes. Phylogenetic comparisons suggest that the tna cluster was acquired by intergeneric lateral transfer, either by H. influenzae or a recent ancestor, and that E. coli may have acquired its tnaA gene from a related source. Genomes of virulent H. influenzae resemble those of pathogenic enterics in having an island of laterally transferred DNA next to mutS. PMID:9422600

  9. Gene-transfer study approval awaits more data

    SciTech Connect

    Marwick, C.

    1988-11-18

    Approval of the gene-transfer study in cancer patients has been delayed. The proposal was recommended for approval by a National Institutes of Health (NIH) advisory committee, but has been put on hold by James B. Wyngaarden, MD, NIH director, pending submission in writing of further information. Some of this information, now forthcoming, had been withheld because data on preliminary studies had been submitted to peer-reviewed journals. The study involves placing the gene for neomycin-resistance to tumor-infiltrating lymphocytes as a marker. When these cells are injected into the patient, the presence of the marker should enable their fate to be studied over a prolonged period and an improved antitumor regimen could result. The use of tumor-infiltrating lymphocytes as immunotherapy has been studied for two years at the NIH's National Cancer Institute. The patients' tumors are removed and the tumor-infiltrating lymphocytes are cultivated to obtain several billion cells. These cells are then injected back into the patient. Early clinical experience has shown a substantial decease in tumor size in some patients, but not in all, an no one knows why.

  10. Adenoviral-mediated gene transfer to rabbit synovium in vivo.

    PubMed Central

    Roessler, B J; Allen, E D; Wilson, J M; Hartman, J W; Davidson, B L

    1993-01-01

    Currently, treatment for rheumatoid arthritis and other inflammatory arthropathies is often ineffective in ameliorating the progression of the disease, particularly the invasive destruction of cartilage and bone by rheumatoid synovium. Multiple aspects of this inflammatory process are mediated by the synovial lining cells (synoviocytes). Genetic modification of these cells in vivo represents a potential method for the treatment of these conditions. In this report, we describe a novel technique for the genetic transduction of synovial lining cells in vivo using recombinant adenoviral vectors and intraarticular injection techniques. Purified high titer suspensions of a recombinant adenoviral vector containing the gene for Escherichia coli beta-galactosidase (AdCMVlacZ) were directly injected into the hind knees of New Zealand white rabbits. Synovial tissues were then examined for transgenic lacZ expression using a combination of in situ staining for beta-galactosidase activity, immunohistochemical staining, and transmission electron microscopy. High efficiency gene transfer and lacZ expression was observed in both type A and type B synoviocytes throughout the articular and periarticular synovium of the rabbit knee, with continued expression of transgenic lacZ detected for > or = 8 wk after infection. Images PMID:8349791

  11. Passive Immunization against HIV/AIDS by Antibody Gene Transfer

    PubMed Central

    Yang, Lili; Wang, Pin

    2014-01-01

    Despite tremendous efforts over the course of many years, the quest for an effective HIV vaccine by the classical method of active immunization remains largely elusive. However, two recent studies in mice and macaques have now demonstrated a new strategy designated as Vectored ImmunoProphylaxis (VIP), which involves passive immunization by viral vector-mediated delivery of genes encoding broadly neutralizing antibodies (bnAbs) for in vivo expression. Robust protection against virus infection was observed in preclinical settings when animals were given VIP to express monoclonal neutralizing antibodies. This unorthodox approach raises new promise for combating the ongoing global HIV pandemic. In this article, we survey the status of antibody gene transfer, review the revolutionary progress on isolation of extremely bnAbs, detail VIP experiments against HIV and its related virus conduced in humanized mice and macaque monkeys, and discuss the pros and cons of VIP and its opportunities and challenges towards clinical applications to control HIV/AIDS endemics. PMID:24473340

  12. Passive immunization against HIV/AIDS by antibody gene transfer.

    PubMed

    Yang, Lili; Wang, Pin

    2014-02-01

    Despite tremendous efforts over the course of many years, the quest for an effective HIV vaccine by the classical method of active immunization remains largely elusive. However, two recent studies in mice and macaques have now demonstrated a new strategy designated as Vectored ImmunoProphylaxis (VIP), which involves passive immunization by viral vector-mediated delivery of genes encoding broadly neutralizing antibodies (bnAbs) for in vivo expression. Robust protection against virus infection was observed in preclinical settings when animals were given VIP to express monoclonal neutralizing antibodies. This unorthodox approach raises new promise for combating the ongoing global HIV pandemic. In this article, we survey the status of antibody gene transfer, review the revolutionary progress on isolation of extremely bnAbs, detail VIP experiments against HIV and its related virus conduced in humanized mice and macaque monkeys, and discuss the pros and cons of VIP and its opportunities and challenges towards clinical applications to control HIV/AIDS endemics. PMID:24473340

  13. Horizontal transference of S-layer genes within Thermus thermophilus.

    PubMed Central

    Fernández-Herrero, L A; Olabarría, G; Castón, J R; Lasa, I; Berenguer, J

    1995-01-01

    The S-layers of Thermus thermophilus HB27 and T. thermophilus HB8 are composed of protein units of 95 kDa (P95) and 100 kDa (P100), respectively. We have selected S-layer deletion mutants from both strains by complete replacement of the slpA gene. Mutants of the two strains showed similar defects in growth and morphology and overproduced an external cell envelope inside of which cells remained after division. However, the nature of this external layer is strain specific, being easily stained and regular in the HB8 delta slpA derivative and amorphous and poorly stained in the HB27 delta slpA strain. The addition of chromosomic DNA from T. thermophilus HB8 to growing cultures of T. thermophilus HB27 delta slpA led to the selection of a new strain, HB27C8, which expressed a functional S-layer composed of the P100 protein. Conversely, the addition of chromosomic DNA from T. thermophilus HB27 to growing cultures of T. thermophilus HB8 delta slpA allowed the isolation of strain HB8C27, which expressed a functional S-layer composed of the P95 protein. The driving force which selected the transference of the S-layer genes in these experiments was the difference in growth rates, one of the main factors leading to selection in natural environments. PMID:7559330

  14. Genome-scale phylogenetic analysis finds extensive gene transfer among fungi

    PubMed Central

    Szöllősi, Gergely J.; Davín, Adrián Arellano; Tannier, Eric; Daubin, Vincent; Boussau, Bastien

    2015-01-01

    Although the role of lateral gene transfer is well recognized in the evolution of bacteria, it is generally assumed that it has had less influence among eukaryotes. To explore this hypothesis, we compare the dynamics of genome evolution in two groups of organisms: cyanobacteria and fungi. Ancestral genomes are inferred in both clades using two types of methods: first, Count, a gene tree unaware method that models gene duplications, gains and losses to explain the observed numbers of genes present in a genome; second, ALE, a more recent gene tree-aware method that reconciles gene trees with a species tree using a model of gene duplication, loss and transfer. We compare their merits and their ability to quantify the role of transfers, and assess the impact of taxonomic sampling on their inferences. We present what we believe is compelling evidence that gene transfer plays a significant role in the evolution of fungi. PMID:26323765

  15. Frequent, independent transfers of a catabolic gene from bacteria to contrasted filamentous eukaryotes

    PubMed Central

    Bruto, Maxime; Prigent-Combaret, Claire; Luis, Patricia; Moënne-Loccoz, Yvan; Muller, Daniel

    2014-01-01

    Even genetically distant prokaryotes can exchange genes between them, and these horizontal gene transfer events play a central role in adaptation and evolution. While this was long thought to be restricted to prokaryotes, certain eukaryotes have acquired genes of bacterial origin. However, gene acquisitions in eukaryotes are thought to be much less important in magnitude than in prokaryotes. Here, we describe the complex evolutionary history of a bacterial catabolic gene that has been transferred repeatedly from different bacterial phyla to stramenopiles and fungi. Indeed, phylogenomic analysis pointed to multiple acquisitions of the gene in these filamentous eukaryotes—as many as 15 different events for 65 microeukaryotes. Furthermore, once transferred, this gene acquired introns and was found expressed in mRNA databases for most recipients. Our results show that effective inter-domain transfers and subsequent adaptation of a prokaryotic gene in eukaryotic cells can happen at an unprecedented magnitude. PMID:24990676

  16. Horizontal gene transfer and gene dosage drives adaptation to wood colonization in a tree pathogen

    PubMed Central

    Dhillon, Braham; Feau, Nicolas; Aerts, Andrea L.; Beauseigle, Stéphanie; Bernier, Louis; Copeland, Alex; Foster, Adam; Gill, Navdeep; Henrissat, Bernard; Herath, Padmini; LaButti, Kurt M.; Levasseur, Anthony; Lindquist, Erika A.; Majoor, Eline; Ohm, Robin A.; Pangilinan, Jasmyn L.; Pribowo, Amadeus; Saddler, John N.; Sakalidis, Monique L.; de Vries, Ronald P.; Grigoriev, Igor V.; Goodwin, Stephen B.; Tanguay, Philippe; Hamelin, Richard C.

    2015-01-01

    Some of the most damaging tree pathogens can attack woody stems, causing lesions (cankers) that may be lethal. To identify the genomic determinants of wood colonization leading to canker formation, we sequenced the genomes of the poplar canker pathogen, Mycosphaerella populorum, and the closely related poplar leaf pathogen, M. populicola. A secondary metabolite cluster unique to M. populorum is fully activated following induction by poplar wood and leaves. In addition, genes encoding hemicellulose-degrading enzymes, peptidases, and metabolite transporters were more abundant and were up-regulated in M. populorum growing on poplar wood-chip medium compared with M. populicola. The secondary gene cluster and several of the carbohydrate degradation genes have the signature of horizontal transfer from ascomycete fungi associated with wood decay and from prokaryotes. Acquisition and maintenance of the gene battery necessary for growth in woody tissues and gene dosage resulting in gene expression reconfiguration appear to be responsible for the adaptation of M. populorum to infect, colonize, and cause mortality on poplar woody stems. PMID:25733908

  17. Towards liver-directed gene therapy: retrovirus-mediated gene transfer into human hepatocytes.

    PubMed

    Grossman, M; Raper, S E; Wilson, J M

    1991-11-01

    Liver-directed gene therapy is being considered in the treatment of inherited metabolic diseases. One approach we are considering is the transplantation of autologous hepatocytes that have been genetically modified with recombinant retroviruses ex vivo. We describe, in this report, techniques for isolating human hepatocytes and efficiently transducing recombinant genes into primary cultures. Hepatocytes were isolated from tissue of four different donors, plated in primary culture, and exposed to recombinant retroviruses expressing either the LacZ reporter gene or the cDNA for rabbit LDL receptor. The efficiency of gene transfer under optimal conditions, as determined by Southern blot analysis, varied from a maximum of one proviral copy per cell to a minimum of 0.1 proviral copy per cell. Cytochemical assays were used to detect expression of the recombinant derived proteins, E. coli beta-galactosidase and rabbit LDL receptor. Hepatocytes transduced with the LDL receptor gene expressed levels of receptor protein that exceeded the normal endogenous levels. The ability to isolate and genetically modify human hepatocytes, as described in this report, is an important step towards the development of liver-directed gene therapies in humans. PMID:1767337

  18. Horizontal gene transfer and gene dosage drives adaptation to wood colonization in a tree pathogen.

    PubMed

    Dhillon, Braham; Feau, Nicolas; Aerts, Andrea L; Beauseigle, Stéphanie; Bernier, Louis; Copeland, Alex; Foster, Adam; Gill, Navdeep; Henrissat, Bernard; Herath, Padmini; LaButti, Kurt M; Levasseur, Anthony; Lindquist, Erika A; Majoor, Eline; Ohm, Robin A; Pangilinan, Jasmyn L; Pribowo, Amadeus; Saddler, John N; Sakalidis, Monique L; de Vries, Ronald P; Grigoriev, Igor V; Goodwin, Stephen B; Tanguay, Philippe; Hamelin, Richard C

    2015-03-17

    Some of the most damaging tree pathogens can attack woody stems, causing lesions (cankers) that may be lethal. To identify the genomic determinants of wood colonization leading to canker formation, we sequenced the genomes of the poplar canker pathogen, Mycosphaerella populorum, and the closely related poplar leaf pathogen, M. populicola. A secondary metabolite cluster unique to M. populorum is fully activated following induction by poplar wood and leaves. In addition, genes encoding hemicellulose-degrading enzymes, peptidases, and metabolite transporters were more abundant and were up-regulated in M. populorum growing on poplar wood-chip medium compared with M. populicola. The secondary gene cluster and several of the carbohydrate degradation genes have the signature of horizontal transfer from ascomycete fungi associated with wood decay and from prokaryotes. Acquisition and maintenance of the gene battery necessary for growth in woody tissues and gene dosage resulting in gene expression reconfiguration appear to be responsible for the adaptation of M. populorum to infect, colonize, and cause mortality on poplar woody stems. PMID:25733908

  19. The Fusarium graminearum Genome Reveals More Secondary Metabolite Gene Clusters and Hints of Horizontal Gene Transfer

    PubMed Central

    Wong, Philip; Münsterkötter, Martin; Mewes, Hans-Werner; Schmeitzl, Clemens; Varga, Elisabeth; Berthiller, Franz; Adam, Gerhard; Güldener, Ulrich

    2014-01-01

    Fungal secondary metabolite biosynthesis genes are of major interest due to the pharmacological properties of their products (like mycotoxins and antibiotics). The genome of the plant pathogenic fungus Fusarium graminearum codes for a large number of candidate enzymes involved in secondary metabolite biosynthesis. However, the chemical nature of most enzymatic products of proteins encoded by putative secondary metabolism biosynthetic genes is largely unknown. Based on our analysis we present 67 gene clusters with significant enrichment of predicted secondary metabolism related enzymatic functions. 20 gene clusters with unknown metabolites exhibit strong gene expression correlation in planta and presumably play a role in virulence. Furthermore, the identification of conserved and over-represented putative transcription factor binding sites serves as additional evidence for cluster co-regulation. Orthologous cluster search provided insight into the evolution of secondary metabolism clusters. Some clusters are characteristic for the Fusarium phylum while others show evidence of horizontal gene transfer as orthologs can be found in representatives of the Botrytis or Cochliobolus lineage. The presented candidate clusters provide valuable targets for experimental examination. PMID:25333987

  20. Improved efficiency of the walnut somatic embryo gene transfer system.

    PubMed

    McGranahan, G H; Leslie, C A; Uratsu, S L; Dandekar, A M

    1990-01-01

    AnAgrobacterium-mediated gene transfer system which relies on repetitive embryogenesis to regenerate transgenic walnut plants has been made more efficient by using a more virulent strain ofAgrobacterium and vectors containing genes for both kanamycin resistance and beta-glucuronidase (GUS) activity to facilitate early screening and selection. Two plasmids (pCGN7001 and pCGN7314) introduced individually into the disarmedAgrobacterium host strain EHA101 were used as inoculum. Embryos maintained on medium containing 100 mg/l kanamycin after co-cultivation produced more transformed secondary embryos than embryos maintained on kanamycin-free medium. Of the 186 GUS-positive secondary embryo lines identified, 70% were regenerated from 3 out of 16 primary embryos inoculated with EHA101/pCGN7314 and grown on kanamycin- containing medium, 28% from 4 out of 17 primary embryos inoculated with EHA101/ pCGN7001 and grown on kanamycin medium, and 2% from one out of 13 primary embryos inoculated with EHA101/pCGN7001 but not exposed to kanamycin. Because kanamycin inhibits but does not completely block new embryo formation in controls, identification of transformants formerly required repetitive selection on kanamycin for several months. Introduction of the GUS marker gene allowed positive identification of transformant secondary embryos as early as 5-6 weeks after inoculation. DNA analysis of a representative subset of lines (n=13) derived from secondary embryos confirmed transformation and provided evidence for multiple insertion events in single inoculated primary embryos. PMID:24226275

  1. Multimodality Imaging of Gene Transfer with a Receptor-Based Reporter Gene

    PubMed Central

    Chen, Ron; Parry, Jesse J.; Akers, Walter J.; Berezin, Mikhail Y.; El Naqa, Issam M.; Achilefu, Samuel; Edwards, W. Barry; Rogers, Buck E.

    2010-01-01

    Gene therapy trials have traditionally used tumor and tissue biopsies for assessing the efficacy of gene transfer. Non-invasive imaging techniques offer a distinct advantage over tissue biopsies in that the magnitude and duration of gene transfer can be monitored repeatedly. Human somatostatin receptor subtype 2 (SSTR2) has been used for the nuclear imaging of gene transfer. To extend this concept, we have developed a somatostatin receptor–enhanced green fluorescent protein fusion construct (SSTR2-EGFP) for nuclear and fluorescent multimodality imaging. Methods An adenovirus containing SSTR2-EGFP (AdSSTR2-EGFP) was constructed and evaluated in vitro and in vivo. SCC-9 human squamous cell carcinoma cells were infected with AdEGFP, AdSSTR2, or AdSSTR2-EGFP for in vitro evaluation by saturation binding, internalization, and fluorescence spectroscopy assays. In vivo biodistribution and nano-SPECT imaging studies were conducted with mice bearing SCC-9 tumor xenografts directly injected with AdSSTR2-EGFP or AdSSTR2 to determine the tumor localization of 111In-diethylenetriaminepentaacetic acid (DTPA)-Tyr3-octreotate. Fluorescence imaging was conducted in vivo with mice receiving intratumoral injections of AdSSTR2, AdSSTR2-EGFP, or AdEGFP as well as ex vivo with tissues extracted from mice. Results The similarity between AdSSTR2-EGFP and wild-type AdSSTR2 was demonstrated in vitro by the saturation binding and internalization assays, and the fluorescence emission spectra of cells infected with AdSSTR2-EGFP was almost identical to the spectra of cells infected with wild-type AdEGFP. Biodistribution studies demonstrated that the tumor uptake of 111In-DTPA-Tyr3-octreotate was not significantly different (P > 0.05) when tumors (n = 5) were injected with AdSSTR2 or AdSSTR2-EGFP but was significantly greater than the uptake in control tumors. Fluorescence was observed in tumors injected with AdSSTR2-EGFP and AdEGFP in vivo and ex vivo but not in tumors injected with AdSSTR2

  2. Persistent Gene Expression in Mouse Nasal Epithelia following Feline Immunodeficiency Virus-Based Vector Gene Transfer

    PubMed Central

    Sinn, Patrick L.; Burnight, Erin R.; Hickey, Melissa A.; Blissard, Gary W.; McCray, Paul B.

    2005-01-01

    Gene transfer development for treatment or prevention of cystic fibrosis lung disease has been limited by the inability of vectors to efficiently and persistently transduce airway epithelia. Influenza A is an enveloped virus with natural lung tropism; however, pseudotyping feline immunodeficiency virus (FIV)-based lentiviral vector with the hemagglutinin envelope protein proved unsuccessful. Conversely, pseudotyping FIV with the envelope protein from influenza D (Thogoto virus GP75) resulted in titers of 106 transducing units (TU)/ml and conferred apical entry into well-differentiated human airway epithelial cells. Baculovirus GP64 envelope glycoproteins share sequence identity with influenza D GP75 envelope glycoproteins. Pseudotyping FIV with GP64 from three species of baculovirus resulted in titers of 107 to 109 TU/ml. Of note, GP64 from Autographa californica multicapsid nucleopolyhedrovirus resulted in high-titer FIV preparations (∼109 TU/ml) and conferred apical entry into polarized primary cultures of human airway epithelia. Using a luciferase reporter gene and bioluminescence imaging, we observed persistent gene expression from in vivo gene transfer in the mouse nose with A. californica GP64-pseudotyped FIV (AcGP64-FIV). Longitudinal bioluminescence analysis documented persistent expression in nasal epithelia for ∼1 year without significant decline. According to histological analysis using a LacZ reporter gene, olfactory and respiratory epithelial cells were transduced. In addition, methylcellulose-formulated AcGP64-FIV transduced mouse nasal epithelia with much greater efficiency than similarly formulated vesicular stomatitis virus glycoprotein-pseudotyped FIV. These data suggest that AcGP64-FIV efficiently transduces and persistently expresses a transgene in nasal epithelia in the absence of agents that disrupt the cellular tight junction integrity. PMID:16188984

  3. Horizontal gene transfer of a Chlamydial tRNA-guanine transglycosylase gene to eukaryotic microbes.

    PubMed

    Manna, Sam; Harman, Ashley

    2016-01-01

    tRNA-guanine transglycosylases are found in all domains of life and mediate the base exchange of guanine with queuine in the anticodon loop of tRNAs. They can also regulate virulence in bacteria such as Shigella flexneri, which has prompted the development of drugs that inhibit the function of these enzymes. Here we report a group of tRNA-guanine transglycosylases in eukaryotic microbes (algae and protozoa) which are more similar to their bacterial counterparts than previously characterized eukaryotic tRNA-guanine transglycosylases. We provide evidence demonstrating that the genes encoding these enzymes were acquired by these eukaryotic lineages via horizontal gene transfer from the Chlamydiae group of bacteria. Given that the S. flexneri tRNA-guanine transglycosylase can be targeted by drugs, we propose that the bacterial-like tRNA-guanine transglycosylases could potentially be targeted in a similar fashion in pathogenic amoebae that possess these enzymes such as Acanthamoeba castellanii. This work also presents ancient prokaryote-to-eukaryote horizontal gene transfer events as an untapped resource of potential drug target identification in pathogenic eukaryotes. PMID:26435002

  4. Transduction-like gene transfer in the methanogen Methanococcus voltae

    NASA Technical Reports Server (NTRS)

    Bertani, G.

    1999-01-01

    Strain PS of Methanococcus voltae (a methanogenic, anaerobic archaebacterium) was shown to generate spontaneously 4.4-kbp chromosomal DNA fragments that are fully protected from DNase and that, upon contact with a cell, transform it genetically. This activity, here called VTA (voltae transfer agent), affects all markers tested: three different auxotrophies (histidine, purine, and cobalamin) and resistance to BES (2-bromoethanesulfonate, an inhibitor of methanogenesis). VTA was most effectively prepared by culture filtration. This process disrupted a fraction of the M. voltae cells (which have only an S-layer covering their cytoplasmic membrane). VTA was rapidly inactivated upon storage. VTA particles were present in cultures at concentrations of approximately two per cell. Gene transfer activity varied from a minimum of 2 x 10(-5) (BES resistance) to a maximum of 10(-3) (histidine independence) per donor cell. Very little VTA was found free in culture supernatants. The phenomenon is functionally similar to generalized transduction, but there is no evidence, for the time being, of intrinsically viral (i.e., containing a complete viral genome) particles. Consideration of VTA DNA size makes the existence of such viral particles unlikely. If they exist, they must be relatively few in number;perhaps they differ from VTA particles in size and other properties and thus escaped detection. Digestion of VTA DNA with the AluI restriction enzyme suggests that it is a random sample of the bacterial DNA, except for a 0.9-kbp sequence which is amplified relative to the rest of the bacterial chromosome. A VTA-sized DNA fraction was demonstrated in a few other isolates of M. voltae.

  5. Transduction-Like Gene Transfer in the Methanogen Methanococcus voltae

    PubMed Central

    Bertani, Giuseppe

    1999-01-01

    Strain PS of Methanococcus voltae (a methanogenic, anaerobic archaebacterium) was shown to generate spontaneously 4.4-kbp chromosomal DNA fragments that are fully protected from DNase and that, upon contact with a cell, transform it genetically. This activity, here called VTA (voltae transfer agent), affects all markers tested: three different auxotrophies (histidine, purine, and cobalamin) and resistance to BES (2-bromoethanesulfonate, an inhibitor of methanogenesis). VTA was most effectively prepared by culture filtration. This process disrupted a fraction of the M. voltae cells (which have only an S-layer covering their cytoplasmic membrane). VTA was rapidly inactivated upon storage. VTA particles were present in cultures at concentrations of approximately two per cell. Gene transfer activity varied from a minimum of 2 × 10−5 (BES resistance) to a maximum of 10−3 (histidine independence) per donor cell. Very little VTA was found free in culture supernatants. The phenomenon is functionally similar to generalized transduction, but there is no evidence, for the time being, of intrinsically viral (i.e., containing a complete viral genome) particles. Consideration of VTA DNA size makes the existence of such viral particles unlikely. If they exist, they must be relatively few in number;perhaps they differ from VTA particles in size and other properties and thus escaped detection. Digestion of VTA DNA with the AluI restriction enzyme suggests that it is a random sample of the bacterial DNA, except for a 0.9-kbp sequence which is amplified relative to the rest of the bacterial chromosome. A VTA-sized DNA fraction was demonstrated in a few other isolates of M. voltae. PMID:10321998

  6. Reduced Serum Level of Interleukin-10 is Associated with Cerebral Infarction: A Case-Control and Meta-Analysis Study.

    PubMed

    Zhu, Yifei; Yang, Haiqing; Diao, Zengyan; Li, Yi; Yan, Chuanzhu

    2016-05-01

    IL-10 expression limits inflammation and restricts the size of CNS damage from stroke. In this study, we examined the correlation between cerebral infarction (CI) and serum levels of interleukin-10 (IL-10) using a combination of case-control study and meta-analysis of published data, with an aim of understanding the relevance of serum IL-10 levels to CI development. This study enrolled a total of 169 CI patients admitted to the Second Hospital of Hebei Medical University between May 2011 and November 2014. During the same period, a group of 145 individuals were recruited at the same hospital as healthy controls after thorough physical examination. Serum IL-10 levels were measured by enzyme-linked immunosorbent assay (ELISA). SPSS 19.0 (IBM, 2010, Chicago, IL, USA) and Comprehensive Meta-Analysis 2.0 (CMA 2.0) software were used for data analysis. Serum levels of IL-10 (pg/mL) were significantly lower in CI patients when compared to healthy controls (15.36 ± 3.21 vs. 21.64 ± 5.17, t = 13.12, P < 0.001). In addition, patients with large artery atherosclerosis (LAAS), cardioembolic infarct (CEI), and lacunar infarct (LAC) displayed drastically reduced serum levels of IL-10 (pg/mL) compared to healthy controls (LAAS 14.77 ± 5.21, CEI 15.25 ± 5.10, LAC 16.58 ± 4.92, all P < 0.001). Interestingly, no significant differences were observed in the serum IL-10 levels when pair-wise comparisons were made between these three clinical subtypes of CI (all P > 0.05). Logistic regression analysis indicated that, with the exception of triglyceride (TG) and uric acid (UA) levels (both P > 0.05), the other seven parameters, including fasting blood glucose (FPG), total cholesterol, low-density lipoprotein (LDL), high-density lipoprotein (HDL), creatinine (Cr), systolic blood pressure (SBP), and diastolic blood pressure (DBP), strongly correlated with CI development (all P < 0.05). Meta-analysis of pooled data from nine case

  7. Calfection: a novel gene transfer method for suspension cells.

    PubMed

    Lindell, Jeanette; Girard, Philippe; Müller, Natalie; Jordan, Martin; Wurm, Florian

    2004-01-20

    We have developed a novel method called Calfection for gene delivery to and protein expression from suspension-cultivated mammalian cells. Plasmid DNA was simply diluted into a calcium chloride solution and then added to the cell culture for transfection. We evaluated and optimized this approach using suspension-adapted HEK293 cells grown in 12-well plates that were shaken on an orbital shaker. Highest expression levels were obtained when cells were transfected at a density of 5x10(5) cells/ml in the presence of 9 mM calcium and 5 microg/ml of plasmid DNA while maintaining a culture pH of 7.6 at the time of transfection. Suspension-adapted BHK 21 and CHO DG 44 cells could also be transfected using this method. Calfection differs from the widely known calcium phosphate coprecipitation technique. The physico-chemical composition of the DNA interacting complexes is not yet known. The transfection cocktail, DNA in a calcium chloride solution, remained highly efficient during long-term storage at temperatures ranging from room temperature to -80 degrees C. In contrast, calcium phosphate-DNA cocktails are only efficient for gene transfer when prepared fresh. Furthermore, passing the calcium-plasmid DNA mixture through a 0.2-microm filter did not compromise protein expression, whereas calcium phosphate-DNA coprecipitates were retained by the filter. High protein expression levels, a limited number of manipulations and the possibility to filter the cocktail make the Calfection approach suitable for both large-scale transfection in bioreactors and for high-throughput transfection experiments in microtiter plates. PMID:14746910

  8. Microbial Evolution Is in the Cards: Horizontal Gene Transfer in the Classroom

    ERIC Educational Resources Information Center

    Kagle, Jeanne; Hay, Anthony G.

    2007-01-01

    Horizontal gene transfer, the exchange of genetic material between bacteria, is a potentially important factor in the degradation of synthetic compounds introduced to the environment and in the acquisition of other characteristics including antibiotic resistance. This game-based activity illustrates the role of horizontal gene transfer in the…

  9. Horizontal gene transfer in the acquisition of novel traits by metazoans

    PubMed Central

    Boto, Luis

    2014-01-01

    Horizontal gene transfer is accepted as an important evolutionary force modulating the evolution of prokaryote genomes. However, it is thought that horizontal gene transfer plays only a minor role in metazoan evolution. In this paper, I critically review the rising evidence on horizontally transferred genes and on the acquisition of novel traits in metazoans. In particular, I discuss suspected examples in sponges, cnidarians, rotifers, nematodes, molluscs and arthropods which suggest that horizontal gene transfer in metazoans is not simply a curiosity. In addition, I stress the scarcity of studies in vertebrates and other animal groups and the importance of forthcoming studies to understand the importance and extent of horizontal gene transfer in animals. PMID:24403327

  10. Gene transfer in the evolution of parasite nucleotide biosynthesis.

    PubMed

    Striepen, Boris; Pruijssers, Andrea J P; Huang, Jinling; Li, Catherine; Gubbels, Marc-Jan; Umejiego, Nwakaso N; Hedstrom, Lizbeth; Kissinger, Jessica C

    2004-03-01

    Nucleotide metabolic pathways provide numerous successful targets for antiparasitic chemotherapy, but the human pathogen Cryptosporidium parvum thus far has proved extraordinarily refractory to classical treatments. Given the importance of this protist as an opportunistic pathogen afflicting immunosuppressed individuals, effective treatments are urgently needed. The genome sequence of C. parvum is approaching completion, and we have used this resource to critically assess nucleotide biosynthesis as a target in C. parvum. Genomic analysis indicates that this parasite is entirely dependent on salvage from the host for its purines and pyrimidines. Metabolic pathway reconstruction and experimental validation in the laboratory further suggest that the loss of pyrimidine de novo synthesis is compensated for by possession of three salvage enzymes. Two of these, uridine kinase-uracil phosphoribosyltransferase and thymidine kinase, are unique to C. parvum within the phylum Apicomplexa. Phylogenetic analysis suggests horizontal gene transfer of thymidine kinase from a proteobacterium. We further show that the purine metabolism in C. parvum follows a highly streamlined pathway. Salvage of adenosine provides C. parvum's sole source of purines. This renders the parasite susceptible to inhibition of inosine monophosphate dehydrogenase, the rate-limiting enzyme in the multistep conversion of AMP to GMP. The inosine 5' monophosphate dehydrogenase inhibitors ribavirin and mycophenolic acid, which are already in clinical use, show pronounced anticryptosporidial activity. Taken together, these data help to explain why widely used drugs fail in the treatment of cryptosporidiosis and suggest more promising targets. PMID:14973196

  11. Neprilysin gene transfer: A promising therapeutic approach for Alzheimer's disease.

    PubMed

    Li, Yuanli; Wang, Junqing; Zhang, Shenghao; Liu, Zhaohui

    2015-09-01

    Alzheimer's disease (AD) is characterized by widespread neurodegeneration throughout the association cortex and limbic system, deposition of amyloid-β (Aβ) in the neuropil and around blood vessels, and formation of neurofibrillary tangles. Aβ accumulation is considered the major pathological change in AD progression. In recent years, several therapeutic strategies for treating AD have focused on reducing the Aβ burden in the brain. Among these approaches, the expression of Aβ-degrading enzymes in the brain has been effective but, so far, impractical for treating patients. Neprilysin (NEP), the most prominent of the Aβ-degrading enzymes in vivo, has been successfully delivered intracranially by viral vectors and is a promising therapeutic approach for reducing Aβ accumulation and treating AD. However, some challenges are associated with the use of viral and nonviral vectors, including secondary toxicity, activation of the immune response, and low efficiency. Therefore, safe and efficient NEP delivery systems that could avoid the viral problems with minor injury and high transfection efficiency are required to deliver AD medical applications. This Mini-Review summarizes NEP gene transfer technologies that use viral and nonviral vectors and discusses the rationale and benefits of these delivery systems for AD treatment trials, providing a reference for basic and clinical studies on AD. PMID:26096375

  12. Gene transfer in the evolution of parasite nucleotide biosynthesis

    PubMed Central

    Striepen, Boris; Pruijssers, Andrea J. P.; Huang, Jinling; Li, Catherine; Gubbels, Marc-Jan; Umejiego, Nwakaso N.; Hedstrom, Lizbeth; Kissinger, Jessica C.

    2004-01-01

    Nucleotide metabolic pathways provide numerous successful targets for antiparasitic chemotherapy, but the human pathogen Cryptosporidium parvum thus far has proved extraordinarily refractory to classical treatments. Given the importance of this protist as an opportunistic pathogen afflicting immunosuppressed individuals, effective treatments are urgently needed. The genome sequence of C. parvum is approaching completion, and we have used this resource to critically assess nucleotide biosynthesis as a target in C. parvum. Genomic analysis indicates that this parasite is entirely dependent on salvage from the host for its purines and pyrimidines. Metabolic pathway reconstruction and experimental validation in the laboratory further suggest that the loss of pyrimidine de novo synthesis is compensated for by possession of three salvage enzymes. Two of these, uridine kinase-uracil phosphoribosyltransferase and thymidine kinase, are unique to C. parvum within the phylum Apicomplexa. Phylogenetic analysis suggests horizontal gene transfer of thymidine kinase from a proteobacterium. We further show that the purine metabolism in C. parvum follows a highly streamlined pathway. Salvage of adenosine provides C. parvum's sole source of purines. This renders the parasite susceptible to inhibition of inosine monophosphate dehydrogenase, the rate-limiting enzyme in the multistep conversion of AMP to GMP. The inosine 5′ monophosphate dehydrogenase inhibitors ribavirin and mycophenolic acid, which are already in clinical use, show pronounced anticryptosporidial activity. Taken together, these data help to explain why widely used drugs fail in the treatment of cryptosporidiosis and suggest more promising targets. PMID:14973196

  13. Identification of the class I genes of the mouse major histocompatibility complex by DNA-mediated gene transfer.

    PubMed

    Goodenow, R S; McMillan, M; Nicolson, M; Sher, B T; Eakle, K; Davidson, N; Hood, L

    1982-11-18

    DNA-mediated gene transfer was used to identify cloned class I genes from the major histocompatibility complex of the BALB/c mouse. Three genes encoding the transplantation antigens H-2 Kd, Dd and Ld were identified as well as genes encoding the Qa-2,3 and two TL differentiation antigens. As many as 10 putative novel class I genes were detected by the association of their gene products with beta 2-microglobulin. Alloantiserum prepared to one of the novel antigens was used to demonstrate the expression of the previously undetected antigen on spleen cells of various inbred, congeneic, and recombinant congeneic strains of mice. PMID:6815535

  14. The Effect of Turmeric (Curcuma longa) Extract on the Functionality of the Solute Carrier Protein 22 A4 (SLC22A4) and Interleukin-10 (IL-10) Variants Associated with Inflammatory Bowel Disease

    PubMed Central

    McCann, Mark J.; Johnston, Sarah; Reilly, Kerri; Men, Xuejing; Burgess, Elaine J.; Perry, Nigel B.; Roy, Nicole C.

    2014-01-01

    Inflammatory bowel disease (IBD) is a chronic relapsing disease. Genetic predisposition to the disease reduces an individual’s capacity to respond appropriately to environmental challenges in the intestine leading to inappropriate inflammation. IBD patients often modify their diet to mitigate or reduce the severity of inflammation. Turmeric (Curcuma longa L., Zingiberaceae) has historically been used in Chinese, Hindu, and Ayurvedic medicine over several centuries to treat inflammatory disorders. To understand how turmeric may influence the consequences of a genetic predisposition to inappropriate inflammation, we used HEK293 cells to examine the in vitro capacity of turmeric extract and fractions to affect the functionality of two gene variants, solute carrier protein 22 A4 (SLC22A4, rs1050152) and interleukin-10 (IL-10, rs1800896) associated with IBD. We found that a turmeric extract and several chromatographically separated fractions beneficially affected the variants of SLC22A4 and IL-10 associated with IBD, by reducing inappropriate epithelial cell transport (SLC22A4, 503F) and increasing anti-inflammatory cytokine gene promoter activity (IL-10, −1082A). The effect of turmeric on the IL-10 variant was strongly associated with the curcumin content of the extract and its fractions. PMID:25314644

  15. Murine AIDS Protects Mice Against Experimental Cerebral Malaria: Down-Regulation by Interleukin 10 a T-Helper Type 1 CD4^+ Cell-Mediated Pathology

    NASA Astrophysics Data System (ADS)

    Eckwalanga, Michel; Marussig, Myriam; Dias Tavares, Marisa; Bouanga, Jean Claude; Hulier, Elisabeth; Henriette Pavlovitch, Jana; Minoprio, Paola; Portnoi, Denis; Renia, Laurent; Mazier, Dominique

    1994-08-01

    The retrovirus LP-BM5 murine leukemia virus induces murine AIDS in C57BL/6 mice that has many similarities with human AIDS; Plasmodium berghei ANKA causes experimental cerebral malaria in the same strain of mice. The outcome of malaria infection was studied in mice concurrently infected with the two pathogens. The retrovirus significantly reduced the gravity of the neurological manifestations associated with Plasmodium berghei ANKA infection. The protection against experimental cerebral malaria induced by murine AIDS increased with duration of viral infection and, hence, with the severity of the immunodeficiency. Interleukin 10, principally from splenic T cells, was shown to play a crucial role in this protection.

  16. Non-Viral Gene Transfer as a Tool for Studying Transcription Regulation of Xenobiotic Metabolizing Enzymes

    PubMed Central

    Bonamassa, Barbara; Liu, Dexi

    2010-01-01

    Numerous xenobiotic metabolizing enzymes are regulated by nuclear receptors at transcriptional level. The challenge we currently face is to understand how a given nuclear receptor interacts with its xenobiotics, migrates into nucleus, binds to the xenobiotic response element of a target gene, and regulates transcription. Toward this end, new methods have been developed to introduce the nuclear receptor gene into appropriate cells and study its activity in activating reporter gene expression under the control of a promoter containing xenobiotic response elements. The goal of this review is to critically examine the gene transfer methods currently available. We concentrate on the gene transfer mechanism, advantages and limitations of each method when employed for nuclear receptor-mediated gene regulation studies. It is our hope that the information provided highlights the importance of gene transfer in studying the mechanisms by which our body eliminates the potentially harmful substances and maintains the homeostasis. PMID:20713102

  17. Apramycin resistance as a selective marker for gene transfer in mycobacteria.

    PubMed Central

    Paget, E; Davies, J

    1996-01-01

    We have explored the potential of using the apramycin resistance gene as a marker in mycobacterial gene transfer studies. Shuttle plasmids available for both electroporation and conjugation studies have been constructed, and we have successfully validated the use of the apramycin resistance gene as a component of cloning vectors for Mycobacterium smegmatis, M. bovis BCG, and M. tuberculosis. PMID:8892841

  18. Migration and horizontal gene transfer divide microbial genomes into multiple niches.

    PubMed

    Niehus, Rene; Mitri, Sara; Fletcher, Alexander G; Foster, Kevin R

    2015-01-01

    Horizontal gene transfer is central to microbial evolution, because it enables genetic regions to spread horizontally through diverse communities. However, how gene transfer exerts such a strong effect is not understood. Here we develop an eco-evolutionary model and show how genetic transfer, even when rare, can transform the evolution and ecology of microbes. We recapitulate existing models, which suggest that asexual reproduction will overpower horizontal transfer and greatly limit its effects. We then show that allowing immigration completely changes these predictions. With migration, the rates and impacts of horizontal transfer are greatly increased, and transfer is most frequent for loci under positive natural selection. Our analysis explains how ecologically important loci can sweep through competing strains and species. In this way, microbial genomes can evolve to become ecologically diverse where different genomic regions encode for partially overlapping, but distinct, ecologies. Under these conditions ecological species do not exist, because genes, not species, inhabit niches. PMID:26592443

  19. Migration and horizontal gene transfer divide microbial genomes into multiple niches

    PubMed Central

    Niehus, Rene; Mitri, Sara; Fletcher, Alexander G.; Foster, Kevin R.

    2015-01-01

    Horizontal gene transfer is central to microbial evolution, because it enables genetic regions to spread horizontally through diverse communities. However, how gene transfer exerts such a strong effect is not understood. Here we develop an eco-evolutionary model and show how genetic transfer, even when rare, can transform the evolution and ecology of microbes. We recapitulate existing models, which suggest that asexual reproduction will overpower horizontal transfer and greatly limit its effects. We then show that allowing immigration completely changes these predictions. With migration, the rates and impacts of horizontal transfer are greatly increased, and transfer is most frequent for loci under positive natural selection. Our analysis explains how ecologically important loci can sweep through competing strains and species. In this way, microbial genomes can evolve to become ecologically diverse where different genomic regions encode for partially overlapping, but distinct, ecologies. Under these conditions ecological species do not exist, because genes, not species, inhabit niches. PMID:26592443

  20. Proteomic profiling of salivary gland after nonviral gene transfer mediated by conventional plasmids and minicircles

    PubMed Central

    Geguchadze, Ramaz; Wang, Zhimin; Zourelias, Lee; Perez-Riveros, Paola; Edwards, Paul C; Machen, Laurie; Passineau, Michael J

    2014-01-01

    In this study, we compared gene transfer efficiency and host response to ultrasound-assisted, nonviral gene transfer with a conventional plasmid and a minicircle vector in the submandibular salivary glands of mice. Initially, we looked at gene transfer efficiency with equimolar amounts of the plasmid and minicircle vectors, corroborating an earlier report showing that minicircle is more efficient in the context of a physical method of gene transfer. We then sought to characterize the physiological response of the salivary gland to exogenous gene transfer using global proteomic profiling. Somewhat surprisingly, we found that sonoporation alone, without a gene transfer vector present, had virtually no effect on the salivary gland proteome. However, when a plasmid vector was used, we observed profound perturbations of the salivary gland proteome that compared in magnitude to that seen in a previous report after high doses of adeno-associated virus. Finally, we found that gene transfer with a minicircle induces only minor proteomic alterations that were similar to sonoporation alone. Using mass spectrometry, we assigned protein IDs to 218 gel spots that differed between plasmid and minicircle. Bioinformatic analysis of these proteins demonstrated convergence on 68 known protein interaction pathways, most notably those associated with innate immunity, cellular stress, and morphogenesis. PMID:25414909

  1. Horizontal Gene Transfer of Pectinases from Bacteria Preceded the Diversification of Stick and Leaf Insects

    PubMed Central

    Shelomi, Matan; Danchin, Etienne G. J.; Heckel, David; Wipfler, Benjamin; Bradler, Sven; Zhou, Xin; Pauchet, Yannick

    2016-01-01

    Genes acquired by horizontal transfer are increasingly being found in animal genomes. Understanding their origin and evolution requires knowledge about the phylogenetic relationships from both source and recipient organisms. We used RNASeq data and respective assembled transcript libraries to trace the evolutionary history of polygalacturonase (pectinase) genes in stick insects (Phasmatodea). By mapping the distribution of pectinase genes on a Polyneoptera phylogeny, we identified the transfer of pectinase genes from known phasmatodean gut microbes into the genome of an early euphasmatodean ancestor that took place between 60 and 100 million years ago. This transfer preceded the rapid diversification of the suborder, enabling symbiont-free pectinase production that would increase the insects’ digestive efficiency and reduce dependence on microbes. Bacteria-to-insect gene transfer was thought to be uncommon, however the increasing availability of large-scale genomic data may change this prevailing notion. PMID:27210832

  2. Horizontal Gene Transfer of Pectinases from Bacteria Preceded the Diversification of Stick and Leaf Insects.

    PubMed

    Shelomi, Matan; Danchin, Etienne G J; Heckel, David; Wipfler, Benjamin; Bradler, Sven; Zhou, Xin; Pauchet, Yannick

    2016-01-01

    Genes acquired by horizontal transfer are increasingly being found in animal genomes. Understanding their origin and evolution requires knowledge about the phylogenetic relationships from both source and recipient organisms. We used RNASeq data and respective assembled transcript libraries to trace the evolutionary history of polygalacturonase (pectinase) genes in stick insects (Phasmatodea). By mapping the distribution of pectinase genes on a Polyneoptera phylogeny, we identified the transfer of pectinase genes from known phasmatodean gut microbes into the genome of an early euphasmatodean ancestor that took place between 60 and 100 million years ago. This transfer preceded the rapid diversification of the suborder, enabling symbiont-free pectinase production that would increase the insects' digestive efficiency and reduce dependence on microbes. Bacteria-to-insect gene transfer was thought to be uncommon, however the increasing availability of large-scale genomic data may change this prevailing notion. PMID:27210832

  3. Horizontal gene transfer and the evolution of transcriptionalregulation in Escherichia coli

    SciTech Connect

    Price, Morgan N.; Dehal, Paramvir S.; Arkin, Adam P.

    2007-12-20

    Background: Most bacterial genes were acquired by horizontalgene transfer from other bacteria instead of being inherited bycontinuous vertical descent from an ancient ancestor}. To understand howthe regulation of these {acquired} genes evolved, we examined theevolutionary histories of transcription factors and of regulatoryinteractions from the model bacterium Escherichia coli K12. Results:Although most transcription factors have paralogs, these usually arose byhorizontal gene transfer rather than by duplication within the E. colilineage, as previously believed. In general, most neighbor regulators --regulators that are adjacent to genes that they regulate -- were acquiredby horizontal gene transfer, while most global regulators evolvedvertically within the gamma-Proteobacteria. Neighbor regulators wereoften acquired together with the adjacent operon that they regulate, sothe proximity might be maintained by repeated transfers (like "selfishoperons"). Many of the as-yet-uncharacterized (putative) regulators havealso been acquired together with adjacent genes, so we predict that theseare neighbor regulators as well. When we analyzed the histories ofregulatory interactions, we found that the evolution of regulation byduplication was rare, and surprisingly, many of the regulatoryinteractions that are shared between paralogs result from convergentevolution. Another surprise was that horizontally transferred genes aremore likely than other genes to be regulated by multiple regulators, andmost of this complex regulation probably evolved after the transfer.Conclusions: Our results highlight the rapid evolution of niche-specificgene regulation in bacteria.

  4. Role of horizontal gene transfer in the evolution of photosynthetic eukaryotes and their plastids.

    PubMed

    Keeling, Patrick J

    2009-01-01

    Plastids are the organelles derived from a cyanobacterium through endosymbiosis. Unlike mitochondria, plastids are not found in all eukaryotes, but their evolution has an added layer of complexity since plastids have moved between eukaryotic lineages by secondary and tertiary endosymbiotic events. This complex history, together with the genetic integration between plastids and their host, has led to many opportunities for gene flow between phylogenetically distinct lineages. Some intracellular transfers do not lead to a protein functioning in a new environment, but many others do and the protein makeup of many plastids appears to have been influenced by exogenous sources as well. Here, different evolutionary sources and cellular destinations of gene flow that has affected the plastid lineage are reviewed. Most horizontal gene transfer (HGT) affecting the modern plastid has taken place via the host nucleus, in the form of genes for plastid-targeted proteins. The impact of this varies greatly from lineage to lineage, but in some cases such transfers can be as high as one fifth of analyzed genes. More rarely, genes have also been transferred to the plastid genome itself, and plastid genes have also been transferred to other non-plant, non-algal lineages. Overall, the proteome of many plastids has emerged as a mosaic of proteins from many sources, some from within the same cell (e.g., cytosolic genes or genes left over from the replacement of an earlier plastid), some from the plastid of other algal lineages, and some from completely unrelated sources. PMID:19271204

  5. Horizontal gene transfer of an entire metabolic pathway between a eukaryotic alga and its DNA virus.

    PubMed

    Monier, Adam; Pagarete, António; de Vargas, Colomban; Allen, Michael J; Read, Betsy; Claverie, Jean-Michel; Ogata, Hiroyuki

    2009-08-01

    Interactions between viruses and phytoplankton, the main primary producers in the oceans, affect global biogeochemical cycles and climate. Recent studies are increasingly revealing possible cases of gene transfers between cyanobacteria and phages, which might have played significant roles in the evolution of cyanobacteria/phage systems. However, little has been documented about the occurrence of horizontal gene transfer in eukaryotic phytoplankton/virus systems. Here we report phylogenetic evidence for the transfer of seven genes involved in the sphingolipid biosynthesis pathway between the cosmopolitan eukaryotic microalga Emiliania huxleyi and its large DNA virus EhV. PCR assays indicate that these genes are prevalent in E. huxleyi and EhV strains isolated from different geographic locations. Patterns of protein and gene sequence conservation support that these genes are functional in both E. huxleyi and EhV. This is the first clear case of horizontal gene transfer of multiple functionally linked enzymes in a eukaryotic phytoplankton-virus system. We examine arguments for the possible direction of the gene transfer. The virus-to-host direction suggests the existence of ancient viruses that controlled the complex metabolic pathway in order to infect primitive eukaryotic cells. In contrast, the host-to-virus direction suggests that the serial acquisition of genes involved in the same metabolic pathway might have been a strategy for the ancestor of EhVs to stay ahead of their closest relatives in the great evolutionary race for survival. PMID:19451591

  6. Horizontal gene transfer of an entire metabolic pathway between a eukaryotic alga and its DNA virus

    PubMed Central

    Monier, Adam; Pagarete, António; de Vargas, Colomban; Allen, Michael J.; Read, Betsy; Claverie, Jean-Michel; Ogata, Hiroyuki

    2009-01-01

    Interactions between viruses and phytoplankton, the main primary producers in the oceans, affect global biogeochemical cycles and climate. Recent studies are increasingly revealing possible cases of gene transfers between cyanobacteria and phages, which might have played significant roles in the evolution of cyanobacteria/phage systems. However, little has been documented about the occurrence of horizontal gene transfer in eukaryotic phytoplankton/virus systems. Here we report phylogenetic evidence for the transfer of seven genes involved in the sphingolipid biosynthesis pathway between the cosmopolitan eukaryotic microalga Emiliania huxleyi and its large DNA virus EhV. PCR assays indicate that these genes are prevalent in E. huxleyi and EhV strains isolated from different geographic locations. Patterns of protein and gene sequence conservation support that these genes are functional in both E. huxleyi and EhV. This is the first clear case of horizontal gene transfer of multiple functionally linked enzymes in a eukaryotic phytoplankton–virus system. We examine arguments for the possible direction of the gene transfer. The virus-to-host direction suggests the existence of ancient viruses that controlled the complex metabolic pathway in order to infect primitive eukaryotic cells. In contrast, the host-to-virus direction suggests that the serial acquisition of genes involved in the same metabolic pathway might have been a strategy for the ancestor of EhVs to stay ahead of their closest relatives in the great evolutionary race for survival. PMID:19451591

  7. Horizontally transferred genes in the genome of Pacific white shrimp, Litopenaeus vannamei

    PubMed Central

    2013-01-01

    Background In recent years, as the development of next-generation sequencing technology, a growing number of genes have been reported as being horizontally transferred from prokaryotes to eukaryotes, most of them involving arthropods. As a member of the phylum Arthropoda, the Pacific white shrimp Litopenaeus vannamei has to adapt to the complex water environments with various symbiotic or parasitic microorganisms, which provide a platform for horizontal gene transfer (HGT). Results In this study, we analyzed the genome-wide HGT events in L. vannamei. Through homology search and phylogenetic analysis, followed by experimental PCR confirmation, 14 genes with HGT event were identified: 12 of them were transferred from bacteria and two from fungi. Structure analysis of these genes showed that the introns of the two fungi-originated genes were substituted by shrimp DNA fragment, two genes transferred from bacteria had shrimp specific introns inserted in them. Furthermore, around other three bacteria-originated genes, there were three large DNA segments inserted into the shrimp genome. One segment was a transposon that fully transferred, and the other two segments contained only coding regions of bacteria. Functional prediction of these 14 genes showed that 6 of them might be related to energy metabolism, and 4 others related to defense of the organism. Conclusions HGT events from bacteria or fungi were happened in the genome of L. vannamei, and these horizontally transferred genes can be transcribed in shrimp. This is the first time to report the existence of horizontally transferred genes in shrimp. Importantly, most of these genes are exposed to a negative selection pressure and appeared to be functional. PMID:23914989

  8. Ultrasound -Assisted Gene Transfer to Adipose Tissue-Derived Stem/Progenitor Cells (ASCs)

    NASA Astrophysics Data System (ADS)

    Miyamoto, Yoshitaka; Ueno, Hitomi; Hokari, Rei; Yuan, Wenji; Kuno, Shuichi; Kakimoto, Takashi; Enosawa, Shin; Negishi, Yoichi; Yoshinaka, Kiyoshi; Matsumoto, Yoichiro; Chiba, Toshio; Hayashi, Shuji

    2011-09-01

    In recent years, multilineage adipose tissue-derived stem cells (ASCs) have become increasingly attractive as a promising source for cell transplantation and regenerative medicine. Particular interest has been expressed in the potential to make tissue stem cells, such as ASCs and marrow stromal cells (MSCs), differentiate by gene transfection. Gene transfection using highly efficient viral vectors such as adeno- and sendai viruses have been developed for this purpose. Sonoporation, or ultrasound (US)-assisted gene transfer, is an alternative gene manipulation technique which employs the creation of a jet stream by ultrasonic microbubble cavitation. Sonoporation using non-viral vectors is expected to be a much safer, although less efficient, tool for prospective clinical gene therapy. In this report, we assessed the efficacy of the sonoporation technique for gene transfer to ASCs. We isolated and cultured adipocyets from mouse adipose tissue. ASCs that have the potential to differentiate with transformation into adipocytes or osteoblasts were obtained. Using the US-assisted system, plasmid DNA containing beta-galactosidase (beta-Gal) and green fluorescent protein (GFP) genes were transferred to the ASCs. For this purpose, a Sonopore 4000 (NEPAGENE Co.) and a Sonazoid (Daiichi Sankyo Co.) instrument were used in combination. ASCs were subjected to US (3.1 MHz, 50% duty cycle, burst rate 2.0 Hz, intensity 1.2 W/cm2, exposure time 30 sec). We observed that the gene was more efficiently transferred with increased concentrations of plasmid DNA (5-150 μg/mL). However, further optimization of the US parameters is required, as the gene transfer efficiency was still relatively low. In conclusion, we herein demonstrate that a gene can be transferred to ASCs using our US-assisted system. In regenerative medicine, this system might resolve the current issues surrounding the use of viral vectors for gene transfer.

  9. Nanoalumina promotes the horizontal transfer of multiresistance genes mediated by plasmids across genera

    PubMed Central

    Qiu, Zhigang; Yu, Yunmei; Chen, Zhaoli; Jin, Min; Yang, Dong; Zhao, Zuguo; Wang, Jingfeng; Shen, Zhiqiang; Wang, Xinwei; Qian, Di; Huang, Aihua; Zhang, Buchang; Li, Jun-Wen

    2012-01-01

    Antibiotic resistance is a worldwide public health concern. Conjugative transfer between closely related strains or species of bacteria is an important method for the horizontal transfer of multidrug-resistance genes. The extent to which nanomaterials are able to cause an increase in antibiotic resistance by the regulation of the conjugative transfer of antibiotic-resistance genes in bacteria, especially across genera, is still unknown. Here we show that nanomaterials in water can significantly promote the horizontal conjugative transfer of multidrug-resistance genes mediated by the RP4, RK2, and pCF10 plasmids. Nanoalumina can promote the conjugative transfer of the RP4 plasmid from Escherichia coli to Salmonella spp. by up to 200-fold compared with untreated cells. We also explored the mechanisms behind this phenomenon and demonstrate that nanoalumina is able to induce oxidative stress, damage bacterial cell membranes, enhance the expression of mating pair formation genes and DNA transfer and replication genes, and depress the expression of global regulatory genes that regulate the conjugative transfer of RP4. These findings are important in assessing the risk of nanomaterials to the environment, particularly from water and wastewater treatment systems, and in the estimation of the effect of manufacture and use of nanomaterials on the environment. PMID:22411796

  10. Gene transfer into experimental brain tumors mediated by adenovirus, herpes simplex virus, and retrovirus vectors.

    PubMed

    Boviatsis, E J; Chase, M; Wei, M X; Tamiya, T; Hurford, R K; Kowall, N W; Tepper, R I; Breakefield, X O; Chiocca, E A

    1994-02-01

    Three vectors derived from retrovirus, herpes simplex virus type 1 (HSV), and adenovirus were compared in cultured rat 9L gliosarcoma cells for gene transfer efficiency and in a 9L rat brain tumor model for histologic pattern and distribution of foreign gene delivery, as well as for associated tumor necrosis and inflammation. At a multiplicity of infection of 1, in vitro transfer of a foreign gene (lacZ from Escherichia coli) into cells was more efficient with either the replication-defective retrovirus vector or the replication-conditional thymidine kinase (TK)-deficient HSV vector than with the replication-defective adenovirus vector. In vivo, stereotactic injections of each vector into rat brain tumors revealed three main histopathologic findings: (i) retrovirus and HSV vector-mediated gene transfer was relatively selective for cells within the tumor, whereas adenovirus vector-mediated gene transfer occurred into several types of endogenous neural cells, as well as into cells within the tumor; (ii) gene transfer to multiple infiltrating tumor deposits without apparent gene transfer to intervening normal brain tissue occurred uniquely in one animal inoculated with the HSV vector, and (iii) extensive necrosis and selective inflammation in the tumor were evident with the HSV vector, whereas there was minimal evidence of tumor necrosis and inflammation with either the retrovirus or adenovirus vectors. PMID:8186298

  11. Indirect Fitness Benefits Enable the Spread of Host Genes Promoting Costly Transfer of Beneficial Plasmids

    PubMed Central

    Dimitriu, Tatiana; Misevic, Dusan; Lotton, Chantal; Brown, Sam P.; Lindner, Ariel B.; Taddei, François

    2016-01-01

    Bacterial genes that confer crucial phenotypes, such as antibiotic resistance, can spread horizontally by residing on mobile genetic elements (MGEs). Although many mobile genes provide strong benefits to their hosts, the fitness consequences of the process of transfer itself are less clear. In previous studies, transfer has been interpreted as a parasitic trait of the MGEs because of its costs to the host but also as a trait benefiting host populations through the sharing of a common gene pool. Here, we show that costly donation is an altruistic act when it spreads beneficial MGEs favoured when it increases the inclusive fitness of donor ability alleles. We show mathematically that donor ability can be selected when relatedness at the locus modulating transfer is sufficiently high between donor and recipients, ensuring high frequency of transfer between cells sharing donor alleles. We further experimentally demonstrate that either population structure or discrimination in transfer can increase relatedness to a level selecting for chromosomal transfer alleles. Both mechanisms are likely to occur in natural environments. The simple process of strong dilution can create sufficient population structure to select for donor ability. Another mechanism observed in natural isolates, discrimination in transfer, can emerge through coselection of transfer and discrimination alleles. Our work shows that horizontal gene transfer in bacteria can be promoted by bacterial hosts themselves and not only by MGEs. In the longer term, the success of cells bearing beneficial MGEs combined with biased transfer leads to an association between high donor ability, discrimination, and mobile beneficial genes. However, in conditions that do not select for altruism, host bacteria promoting transfer are outcompeted by hosts with lower transfer rate, an aspect that could be relevant in the fight against the spread of antibiotic resistance. PMID:27270455

  12. Biomaterial-Mediated Retroviral Gene Transfer Using Self-Assembled Monolayers

    PubMed Central

    Gersbach, Charles A.; Coyer, Sean R.; Le Doux, Joseph M.; García, Andrés J.

    2007-01-01

    Biomaterial-mediated gene delivery has recently emerged as a promising alternative to conventional gene transfer technologies that focus on direct delivery of viral vectors or DNA-polymer/matrix complexes. However, biomaterial-based strategies have primarily targeted transient gene expression vehicles, including plasmid DNA and adenovirus particles. This study expands on this work by characterizing biomaterial properties conducive to the surface immobilization of retroviral particles and subsequent transduction of mammalian cells at the cell-material interface. Self-assembled monolayers (SAMs) of functionally-terminated alkanethiols on gold were used to establish biomaterial surfaces of defined chemical composition. Gene transfer was observed to be greater than 90% on NH2-terminated surfaces, approximately 50% on COOH-functionalized surfaces, and undetectable on CH3-terminated SAMs, similar to controls of tissue culture-treated polystyrene. Gene delivery via the NH2-SAM was further characterized as a function of coating time, virus concentration, and cell seeding density. Finally, SAM-mediated gene delivery was comparable to fibronectin- and poly-L-lysine-based methods for gene transfer. This work is significant to establishing safe and effective gene therapy strategies, developing efficient methods for gene delivery, and supporting recent progress in the field of biomaterial-mediated gene transfer. PMID:17698189

  13. An ancient horizontal gene transfer between mosquito and the endosymbiotic bacterium Wolbachia pipientis.

    PubMed

    Woolfit, Megan; Iturbe-Ormaetxe, Iñaki; McGraw, Elizabeth A; O'Neill, Scott L

    2009-02-01

    The extent and biological relevance of horizontal gene transfer (HGT) in eukaryotic evolution remain highly controversial. Recent studies have demonstrated frequent and large-scale HGT from endosymbiotic bacteria to their hosts, but the great majority of these transferred genes rapidly become nonfunctional in the recipient genome. Here, we investigate an ancient HGT between a host metazoan and an endosymbiotic bacterium, Wolbachia pipientis. The transferred gene has so far been found only in mosquitoes and Wolbachia. In mosquitoes, it is a member of a gene family encoding candidate receptors required for malaria sporozoite invasion of the mosquito salivary gland. The gene copy in Wolbachia has substantially diverged in sequence from the mosquito homolog, is evolving under purifying selection, and is expressed, suggesting that this gene is also functional in the bacterial genome. Several lines of evidence indicate that the gene may have been transferred from eukaryotic host to bacterial endosymbiont. Regardless of the direction of transfer, however, these results demonstrate that interdomain HGT may give rise to functional, persistent, and possibly evolutionarily significant new genes. PMID:18988686

  14. SAFETY AND EFFICIENCY OF MODULATING PARACELLULAR PERMEABILITY TO ENHANCE AIRWAY EPITHELIAL GENE TRANSFER IN VIVO

    EPA Science Inventory


    ABSTRACT

    We evaluated the safety of agents that enhance gene transfer by modulating paracellular permeability. Lactate dehydrogenase (LDH) and cytokine release were measured in polarized primary human airway epithelial (HAE) cells after luminal application of vehicle, ...

  15. An XMRV Derived Retroviral Vector as a Tool for Gene Transfer

    PubMed Central

    2011-01-01

    Background Retroviral vectors are widely used tools for gene delivery and gene therapy. They are useful for gene expression studies and genetic manipulation in vitro and in vivo. Many retroviral vectors are derived from the mouse gammaretrovirus, murine leukemia virus (MLV). These vectors have been widely used in gene therapy clinical trials. XMRV, initially found in prostate cancer tissue, was the first human gammaretrovirus described. Findings We developed a new retroviral vector based on XMRV called pXC. It was developed for gene transfer to human cells and is produced by transient cotransfection of LNCaP cells with pXC and XMRV-packaging plasmids. Conclusions We demonstrated that pXC mediates expression of inserted transgenes in cell lines. This new vector will be a useful tool for gene transfer in human and non-human cell lines, including gene therapy studies. PMID:21651801

  16. Evolutionary Advantage Conferred by an Eukaryote-to-Eukaryote Gene Transfer Event in Wine Yeasts

    PubMed Central

    Marsit, Souhir; Mena, Adriana; Bigey, Frédéric; Sauvage, François-Xavier; Couloux, Arnaud; Guy, Julie; Legras, Jean-Luc; Barrio, Eladio; Dequin, Sylvie; Galeote, Virginie

    2015-01-01

    Although an increasing number of horizontal gene transfers have been reported in eukaryotes, experimental evidence for their adaptive value is lacking. Here, we report the recent transfer of a 158-kb genomic region between Torulaspora microellipsoides and Saccharomyces cerevisiae wine yeasts or closely related strains. This genomic region has undergone several rearrangements in S. cerevisiae strains, including gene loss and gene conversion between two tandemly duplicated FOT genes encoding oligopeptide transporters. We show that FOT genes confer a strong competitive advantage during grape must fermentation by increasing the number and diversity of oligopeptides that yeast can utilize as a source of nitrogen, thereby improving biomass formation, fermentation efficiency, and cell viability. Thus, the acquisition of FOT genes has favored yeast adaptation to the nitrogen-limited wine fermentation environment. This finding indicates that anthropic environments offer substantial ecological opportunity for evolutionary diversification through gene exchange between distant yeast species. PMID:25750179

  17. Direct gene transfer into human cultured cells facilitated by laser micropuncture of the cell membrane

    SciTech Connect

    Tao, W.; Wilkinson, J.; Stanbridge, E.J.; Berns, M.W.

    1987-06-01

    The selective alteration of the cellular genome by laser microbeam irradiation has been extensively applied in cell biology. We report here the use of the third harmonic (355 nm) of an yttrium-aluminum garnet laser to facilitate the direct transfer of the neo gene into cultured human HT1080-6TG cells. The resultant transformants were selected in media containing an aminoglycoside antibiotic, G418. Integration of the neo gene into individual chromosomes and expression of the gene were demonstrated by Southern blot analyses, microcell-mediated chromosome transfer, and chromosome analyses. The stability of the integrated neo gene in the transformants was shown by a comparative growth assay in selective and nonselective media. Transformation and incorporation of the neo gene into the host genome occurred at a frequency of 8x10-4-3x10-3. This method appears to be 100-fold more efficient than the standard calcium phosphate-mediated method of DNA transfer.

  18. Direct Gene Transfer into Human Cultured Cells Facilitated by Laser Micropuncture of the Cell Membrane

    NASA Astrophysics Data System (ADS)

    Tao, Wen; Wilkinson, Joyce; Stanbridge, Eric J.; Berns, Michael W.

    1987-06-01

    The selective alteration of the cellular genome by laser microbeam irradiation has been extensively applied in cell biology. We report here the use of the third harmonic (355 nm) of an yttrium-aluminum garnet laser to facilitate the direct transfer of the neo gene into cultured human HT1080-6TG cells. The resultant transformants were selected in medium containing an aminoglycoside antibiotic, G418. Integration of the neo gene into individual human chromosomes and expression of the gene were demonstrated by Southern blot analyses, microcell-mediated chromosome transfer, and chromosome analyses. The stability of the integrated neo gene in the transformants was shown by a comparative growth assay in selective and nonselective media. Transformation and incorporation of the neo gene into the host genome occurred at a frequency of 8 × 10-4-3 × 10-3. This method appears to be 100-fold more efficient than the standard calcium phosphate-mediated method of DNA transfer.

  19. Quantitative analysis of recombination between YFP and CFP genes of FRET biosensors introduced by lentiviral or retroviral gene transfer.

    PubMed

    Komatsubara, Akira T; Matsuda, Michiyuki; Aoki, Kazuhiro

    2015-01-01

    Biosensors based on the principle of Förster (or fluorescence) resonance energy transfer (FRET) have been developed to visualize spatio-temporal dynamics of signalling molecules in living cells. Many of them adopt a backbone of intramolecular FRET biosensor with a cyan fluorescent protein (CFP) and yellow fluorescent protein (YFP) as donor and acceptor, respectively. However, there remains the difficulty of establishing cells stably expressing FRET biosensors with a YFP and CFP pair by lentiviral or retroviral gene transfer, due to the high incidence of recombination between YFP and CFP genes. To address this, we examined the effects of codon-diversification of YFP on the recombination of FRET biosensors introduced by lentivirus or retrovirus. The YFP gene that was fully codon-optimized to E.coli evaded the recombination in lentiviral or retroviral gene transfer, but the partially codon-diversified YFP did not. Further, the length of spacer between YFP and CFP genes clearly affected recombination efficiency, suggesting that the intramolecular template switching occurred in the reverse-transcription process. The simple mathematical model reproduced the experimental data sufficiently, yielding a recombination rate of 0.002-0.005 per base. Together, these results show that the codon-diversified YFP is a useful tool for expressing FRET biosensors by lentiviral or retroviral gene transfer. PMID:26290434

  20. Development of second- and third-generation bovine immunodeficiency virus-based gene transfer systems.

    PubMed

    Matukonis, Meghan; Li, Mengtao; Molina, Rene P; Paszkiet, Brian; Kaleko, Michael; Luo, Tianci

    2002-07-20

    Lentivirus-based gene transfer systems have demonstrated their utility in mediating gene transfer to dividing and nondividing cells both in vitro and in vivo. An early-generation gene transfer system developed from bovine immunodeficiency virus (BIV) has been described (Berkowitz et al., J. Virol. 2001;75:3371-3382). In this paper, we describe the development of second-generation (three-plasmid) and third-generation (four-plasmid) BIV-based systems. All accessory genes (vif, vpw, vpy, and tmx) and the regulatory gene tat were deleted or largely truncated from the packaging construct. Furthermore, we split the packaging function into two constructs by expressing Rev in a separate plasmid. Together with our minimal BIV transfer vector construct and a vesicular stomatitis virus G glycoprotein-expressing plasmid, the BIV vectors were generated. The vectors produced by the three- and four-plasmid systems had titers greater than 1 x 10(6) transducing units per milliliter and were fully functional as indicated by their ability to efficiently transduce both dividing and nondividing cells. These results suggest that the accessory genes vif, vpw, vpy, and tmx are dispensable for functional BIV vector development. The modifications made to the packaging constructs improve the safety profile of the vector system. Finally, BIV vectors provide an alternative to human immunodeficiency virus-based gene transfer systems. PMID:12162812

  1. Exploration of horizontal gene transfer between transplastomic tobacco and plant-associated bacteria.

    PubMed

    Demanèche, Sandrine; Monier, Jean-Michel; Dugat-Bony, Eric; Simonet, Pascal

    2011-10-01

    The likelihood of gene transfer from transgenic plants to bacteria is dependent on the transgene copy number and on the presence of homologous sequences for recombination. The large number of chloroplast genomes in a plant cell as well as the prokaryotic origin of the transgene may thus significantly increase the likelihood of gene transfer from transplastomic plants to bacteria. In order to assess the probability of such a transfer, bacterial isolates, screened for their ability to colonize decaying tobacco plant tissue and possessing DNA sequence similarity to the chloroplastic genes accD and rbcL flanking the transgene (aadA), were tested for their ability to take up extracellular DNA (broad host-range pBBR1MCS-3-derived plasmid, transplastomic plant DNA and PCR products containing the genes accD-aadA-rbcL) by natural or electrotransformation. The results showed that among the 16 bacterial isolates tested, six were able to accept foreign DNA and acquire the spectinomycin resistance conferred by the aadA gene on plasmid, but none of them managed to integrate transgenic DNA in their chromosome. Our results provide no indication that the theoretical gene transfer-enhancing properties of transplastomic plants cause horizontal gene transfer at rates above those found in other studies with nuclear transgenes. PMID:21564143

  2. Interleukin-10 -1082A/G polymorphism is associated with the development of acute pancreatitis in a Chinese population

    PubMed Central

    Cai, Feng; Cui, Ning; Ma, Hongyan; Wang, Xueli; Qiao, Guihong; Liu, Danping

    2015-01-01

    We conducted a case-control study to investigate the association between IL-10 gene polymorphism (-1082A/G, -819T/C, and -592A/C) and risk of acute pancreatitis in a Chinese population. A total of 240 patients with proven acute pancreatitis and 240 control subjects were collected between May 2012 and January 2015. Genotyping of the IL-10-1082A/G, -819T/C, and -592A/C gene polymorphisms was conducted by using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. By univariate logistic regression analysis, patients with acute pancreatitis were more likely to have higher BMI (OR=2.12, 95% CI=1.45-3.12; P<0.001) and have a habit of alcohol drinking (OR=2.01, 95% CI=1.37-2.95; P<0.001). There were significant differences in the genotype distributions of IL-10-1082A/G between patients with acute pancreatitis and control subjects (χ2=9.97, P=0.007). By multiple logistic regression analysis, we found that individuals with the GG genotype of IL-10-1082A/G were associated with an increased risk of acute pancreatitis when compared with the AA genotype (OR=2.32, 95% CI=1.20-4.59; P=0.007). In dominant and recessive models, the IL-10-1082A/G gene polymorphism was significantly correlated with an elevated risk of acute pancreatitis, and the adjusted Ors (95% CI) were 1.50 (1.03-2.20) and 1.99 (1.06-3.79), respectively. However, no significant different was found between IL-10-819T/C and -592A/C gene polymorphisms and susceptibility to acute pancreatitis. In conclusion, we suggest that IL-10-1082A/G gene polymorphisms contribute to the development of acute pancreatitis in codominant, dominant and recessive models. PMID:26823862

  3. Chloroplast-like transfer RNA genes expressed in wheat mitochondria.

    PubMed Central

    Joyce, P B; Gray, M W

    1989-01-01

    In the course of a systematic survey of wheat mitochondrial tRNA genes, we have sequenced chloroplast-like serine (trnS-GGA), phenylalanine (trnF-GAA) and cysteine (trnC-GCA) tRNA genes and their flanking regions. These genes are remnants of 'promiscuous' chloroplast DNA that has been incorporated into wheat mtDNA in the course of its evolution. Each gene differs by one or a few nucleotides from the authentic chloroplast homolog previously characterized in wheat or other plants, and each could potentially encode a functional tRNA whose secondary structure shows no deviations from the generalized model. To determine whether these chloroplast-like tRNA genes are actually expressed, wheat mitochondrial tRNAs were resolved by a series of polyacrylamide gel electrophoreses, after being specifically end-labeled in vitro by 3'-CCA addition mediated by wheat tRNA nucleotidyltransferase. Subsequent direct RNA sequence analysis identified prominent tRNA species corresponding to the mitochondrial and not the chloroplast trnS, trnF and trnC genes. This analysis also revealed chloroplast-like elongator methionine, asparagine and tryptophan tRNAs. Our results suggest that at least some chloroplast-like tRNA genes in wheat mtDNA are transcribed, with transcripts undergoing processing, post-transcriptional modification and 3'-CCA addition, to produce mature tRNAs that may participate in mitochondrial protein synthesis. Images PMID:2762145

  4. Lentiviral vector-mediated gene transfer and RNA silencing technology in neuronal dysfunctions.

    PubMed

    Dreyer, Jean-Luc

    2011-02-01

    Lentiviral-mediated gene transfer in vivo or in cultured mammalian neurons can be used to address a wide variety of biological questions, to design animals models for specific neurodegenerative pathologies, or to test potential therapeutic approaches in a variety of brain disorders. Lentiviruses can infect non-dividing cells, thereby allowing stable gene transfer in post-mitotic cells such as mature neurons. An important contribution has been the use of inducible vectors: the same animal can thus be used repeatedly in the doxycycline-on or -off state, providing a powerful mean for assessing the function of a gene candidate in a disorder within a specific neuronal circuit. Furthermore, lentivirus vectors provide a unique tool to integrate siRNA expression constructs with the aim to locally knockdown expression of a specific gene, enabling to assess the function of a gene in a very specific neuronal pathway. Lentiviral vector-mediated delivery of short hairpin RNA results in persistent knockdown of gene expression in the brain. Therefore, the use of lentiviruses for stable expression of siRNA in brain is a powerful aid to probe gene functions in vivo and for gene therapy of diseases of the central nervous system. In this chapter I review the applications of lentivirus-mediated gene transfer in the investigation of specific gene candidates involved in major brain disorders and neurodegenerative processes. Major applications have been in polyglutamine disorders, such as synucleinopathies and Parkinson's disease, or in investigating gene function in Huntington's disease, dystonia, or muscular dystrophy. Recently, lentivirus gene transfer has been an invaluable tool for evaluation of gene function in behavioral disorders such as drug addiction and attention-deficit hyperactivity disorder or in learning and cognition. PMID:20862616

  5. Increase of interleukin-10-producing B cells associated with long-term remission after i.v. immunoglobulin treatment for pemphigus.

    PubMed

    Kabuto, Miho; Fujimoto, Noriki; Tanaka, Toshihiro

    2016-07-01

    We present a refractory case of pemphigus vulgaris that achieved long-term remission after i.v. immunoglobulin treatment (IVIG). We evaluated the fluctuation of circulating interleukin-10-producing B cells (B10 cells) during the course in our case and other three patients with pemphigus treated with IVIG without clinical remission. B10 cells were observed predominantly in CD1d(-) , CD5(-) , CD9(-) and CD27(+) populations among CD19(+) cells in healthy controls, as well as in patients with pemphigus. The frequency of B10 cells among CD19(+) cells increased in our case, but not in the other three patients without clinical remission, which leads to speculation on the association between the increase of B10 cells and the achievement of long-term remission after IVIG treatment. PMID:26871259

  6. Gene transfer into hematopoietic progenitor and stem cells: progress and problems.

    PubMed

    Dunbar, C E; Emmons, R V

    1994-11-01

    Gene transfer to hematopoietic cells for the purpose of "gene therapy" is a new and rapidly developing field with clinical trials in progress. A fundamental goal of research in this field is the incorporation of exogenous genes into the chromosomes of the most primitive hematopoietic progenitor cells--stem cells. Recombinantly engineered retroviral vectors are the best characterized and are currently the only vector type in clinical trials directed at the hematopoietic system. High efficiency gene transfer and expression in murine stem cells and their progeny is now routine, but in larger animal models such as dogs or primates and preliminary clinical trials, gene transfer has been less successful. Problems such as retroviral efficiency, gene expression, insertional mutagenesis and helper virus contamination are being addressed. A promising new vector, the adeno-associated virus (AAV), has shown promise and may allow production of high titer, stable, recombinant virions without helper contamination and with potentially better safety parameters. However, the technology for AAV gene transfer is currently underdeveloped, and issues related to the reproducible production of vectors must be addressed. Other non-viral vector systems are being explored, but little data are available on applications to hematopoietic cells. Better preclinical models are needed to study gene targeting and expression in human cells. An overview of recombinant retroviral and adeno-associated viral vector production, preclinical data and preliminary clinical data will be given, and problems needing to be addressed at all stages of development before broad clinical utility can be achieved will be discussed. PMID:7881358

  7. Evolutionary transfer of the chloroplast tufA gene to the nucleus.

    PubMed

    Baldauf, S L; Palmer, J D

    1990-03-15

    Evolutionary gene transfer is a basic corollary of the now widely accepted endosymbiotic theory, which proposes that mitochondria and chloroplasts originated from once free-living eubacteria. The small organellar chromosomes are remnants of larger bacterial genomes, with most endosymbiont genes having been either transferred to the nucleus soon after endosymbiosis or lost entirely, with some being functionally replaced by pre-existing nuclear genes. Several lines of evidence indicate that relocation of some organelle genes could have been more recent. These include the abundance of non-functional organelle sequences of recent origin in nuclear DNA, successful artificial transfer of functional organelle genes to the nucleus, and several examples of recently lost organelle genes, although none of these is known to have been replaced by a nuclear homologue that is clearly of organellar ancestry. We present gene sequence and molecular phylogenetic evidence for the transfer of the chloroplast tufA gene to the nucleus in the green algal ancestor of land plants. PMID:2314461

  8. The combination of IκB kinase β inhibitor and everolimus modulates expression of interleukin-10 in human T-cell lymphotropic virus type-1-infected T cells.

    PubMed

    Nishioka, Chie; Ikezoe, Takayuki; Yang, Jing; Udaka, Keiko; Yokoyama, Akihito

    2013-03-01

    Adult T-cell leukaemia-lymphoma (ATLL) is an aggressive malignancy of CD4(+)  CD25(+) T lymphocytes, characterized by a severely compromised immunosystem, in which the human T-cell lymphotropic virus type 1 (HTLV-1) has been recognized as the aetiological agent. This study found that an IκB kinase β (IKKβ) inhibitor Bay11-7082 inactivated mammalian target of rapamycin (mTOR), signal transducer and activator of transcription 3 and transcription factor nuclear factor-κB in HTLV-1-infected T cells; this was significantly enhanced in the presence of the mTOR inhibitor everolimus. In addition, Bay11-7082 decreased production of the immunosuppressive cytokine interleukin-10 (IL-10), which was further down-regulated when Bay11-7082 was combined with evelolimus in HTLV-1-infected T and ATLL cells isolated from patients. Interleukin-10 is known to inhibit maturation and the antigen-presenting function of dendritic cells (DCs). The culture media of HTLV-1-infected MT-1 cells, which contained a large amout of IL-10, hampered tumour necrosis factor-α-induced maturation of DCs isolated from healthy volunteers. Culture supernatant of MT-1 cells treated with a combination of Bay11-7082 and everolimus augmented maturation of DCs in association with a decrease in production of IL-10 and enhanced the allostimulatory function of DCs. Similarly, when DCs isolated from patients with ATLL were treated with the combination of Bay11-7082 and everolimus, they were fully matured and their capability to stimulate proliferation of lymphocytes was augmented. Taken together, the combination of Bay11-7082 and everolimus might exhibit immunostimulatory properties in HTLV-1-infected T and ATLL cells isolated from patients, and this combination may be potentially therapeutic to regain the compromised immunosystem in ATLL patients. PMID:23278479

  9. Horizontal Gene Transfers from Bacteria to Entamoeba Complex: A Strategy for Dating Events along Species Divergence.

    PubMed

    Romero, Miguel; Cerritos, R; Ximenez, Cecilia

    2016-01-01

    Horizontal gene transfer has proved to be relevant in eukaryotic evolution, as it has been found more often than expected and related to adaptation to certain niches. A relatively large list of laterally transferred genes has been proposed and evaluated for the parasite Entamoeba histolytica. The goals of this work were to elucidate the importance of lateral gene transfer along the evolutionary history of some members of the genus Entamoeba, through identifying donor groups and estimating the divergence time of some of these events. In order to estimate the divergence time of some of the horizontal gene transfer events, the dating of some Entamoeba species was necessary, following an indirect dating strategy based on the fossil record of plausible hosts. The divergence between E. histolytica and E. nuttallii probably occurred 5.93 million years ago (Mya); this lineage diverged from E. dispar 9.97 Mya, while the ancestor of the latter separated from E. invadens 68.18 Mya. We estimated times for 22 transferences; the most recent occurred 31.45 Mya and the oldest 253.59 Mya. Indeed, the acquisition of genes through lateral transfer may have triggered a period of adaptive radiation, thus playing a major role in the evolution of the Entamoeba genus. PMID:27239333

  10. Horizontal Gene Transfers from Bacteria to Entamoeba Complex: A Strategy for Dating Events along Species Divergence

    PubMed Central

    Romero, Miguel; Ximenez, Cecilia

    2016-01-01

    Horizontal gene transfer has proved to be relevant in eukaryotic evolution, as it has been found more often than expected and related to adaptation to certain niches. A relatively large list of laterally transferred genes has been proposed and evaluated for the parasite Entamoeba histolytica. The goals of this work were to elucidate the importance of lateral gene transfer along the evolutionary history of some members of the genus Entamoeba, through identifying donor groups and estimating the divergence time of some of these events. In order to estimate the divergence time of some of the horizontal gene transfer events, the dating of some Entamoeba species was necessary, following an indirect dating strategy based on the fossil record of plausible hosts. The divergence between E. histolytica and E. nuttallii probably occurred 5.93 million years ago (Mya); this lineage diverged from E. dispar 9.97 Mya, while the ancestor of the latter separated from E. invadens 68.18 Mya. We estimated times for 22 transferences; the most recent occurred 31.45 Mya and the oldest 253.59 Mya. Indeed, the acquisition of genes through lateral transfer may have triggered a period of adaptive radiation, thus playing a major role in the evolution of the Entamoeba genus. PMID:27239333

  11. Kinetics of conjugative gene transfer on surfaces in granular porous media

    NASA Astrophysics Data System (ADS)

    Massoudieh, A.; Crain, C.; Lambertini, E.; Nelson, K. E.; Barkouki, T.; L'Amoreaux, P.; Loge, F. J.; Ginn, T. R.

    2010-03-01

    The transfer of genetic material among bacteria in the environment can occur both in the planktonic and attached state. Given the propensity of organisms to exist in sessile microbial communities in oligotrophic subsurface conditions, and that such conditions typify the subsurface, this study focuses on exploratory modeling of horizontal gene transfer among surface-associated Escherichiacoli in the subsurface. The mathematics so far used to describe the kinetics of conjugation in biofilms are developed largely from experimental observations of planktonic gene transfer, and are absent of lags or plasmid stability that appear experimentally. We develop a model and experimental system to quantify bacterial filtration and gene transfer in the attached state, on granular porous media. We include attachment kinetics described in Nelson et al. (2007) using the filtration theory approach of Nelson and Ginn (2001, 2005) with motility of E. coli described according to Biondi et al. (1998).

  12. Kinetics of conjugative gene transfer on surfaces in granular porous media

    NASA Astrophysics Data System (ADS)

    Ginn, T.; Massoudieh, A.; Nelson, K.; Mathew, A.; Lambertini, E.

    2005-12-01

    The transfer of genetic material among bacteria in the environment can occur both in the planktonic and attached state. Given the propensity of organisms to exist in sessile microbial communities in oligotrophic conditions, and that such conditions typify the subsurface, this study focuses on exploratory modeling of horizontal gene transfer among surface-associated E. coli in the subsurface. The mathematics so far used to describe the kinetics of conjugation in biofilms are developed largely from experimental observations of planktonic gene transfer, and are absent of lags or plasmid stability that appear experimentally. We develop a model for bacterial filtration and gene transfer in the attached state, for the early stages of biofilm formation using a recently revised filtration theory approach (Nelson and Ginn, 2005) with motility of E. coli described as a continuous time random walk according to data from microflow chamber experiments (Biondi et al., 2002).

  13. Limitations of the murine nose in the development of nonviral airway gene transfer.

    PubMed

    Griesenbach, Uta; Sumner-Jones, Stephanie G; Holder, Emma; Munkonge, Felix M; Wodehouse, Theresa; Smith, Stephen N; Wasowicz, Marguerite Y; Pringle, Ian; Casamayor, Isabel; Chan, Mario; Coles, Rebecca; Cornish, Nikki; Dewar, Ann; Doherty, Ann; Farley, Raymond; Green, Anne-Marie; Jones, Bryony L; Larsen, Mia D B; Lawton, Anna E; Manvell, Michelle; Painter, Hazel; Singh, Charanjit; Somerton, Lucinda; Stevenson, Barbara; Varathalingam, Anusha; Siegel, Craig; Scheule, Ronald K; Cheng, Seng H; Davies, Jane C; Porteous, David J; Gill, Deborah R; Boyd, A Christopher; Hyde, Steve C; Alton, Eric W F W

    2010-07-01

    A clinical program to assess whether lipid GL67A-mediated gene transfer can ameliorate cystic fibrosis (CF) lung disease is currently being undertaken by the UK CF Gene Therapy Consortium. We have evaluated GL67A gene transfer to the murine nasal epithelium of wild-type and CF knockout mice to assess this tissue as a test site for gene transfer agents. The plasmids used were regulated by either (1) the commonly used short-acting cytomegalovirus promoter/enhancer or (2) the ubiquitin C promoter. In a study of approximately 400 mice with CF, vector-specific CF transmembrane conductance regulator (CFTR) mRNA was detected in nasal epithelial cells of 82% of mice treated with a cytomegalovirus-plasmid (pCF1-CFTR), and 62% of mice treated with an ubiquitin C-plasmid. We then assessed whether CFTR gene transfer corrected a panel of CFTR-specific endpoint assays in the murine nose, including ion transport, periciliary liquid height, and ex vivo bacterial adherence. Importantly, even with the comparatively large number of animals assessed, the CFTR function studies were only powered to detect changes of more than 50% toward wild-type values. Within this limitation, no significant correction of the CF phenotype was detected. At the current levels of gene transfer efficiency achievable with nonviral vectors, the murine nose is of limited value as a stepping stone to human trials. PMID:19648474

  14. Complexity of genetic sequences modified by horizontal gene transfer and degraded-DNA uptake

    NASA Astrophysics Data System (ADS)

    Tremberger, George; Dehipawala, S.; Nguyen, A.; Cheung, E.; Sullivan, R.; Holden, T.; Lieberman, D.; Cheung, T.

    2015-09-01

    Horizontal gene transfer has been a major vehicle for efficient transfer of genetic materials among living species and could be one of the sources for noncoding DNA incorporation into a genome. Our previous study of lnc- RNA sequence complexity in terms of fractal dimension and information entropy shows a tight regulation among the studied genes in numerous diseases. The role of sequence complexity in horizontal transferred genes was investigated with Mealybug in symbiotic relation with a 139K genome microbe and Deinococcus radiodurans as examples. The fractal dimension and entropy showed correlation R-sq of 0.82 (N = 6) for the studied Deinococcus radiodurans sequences. For comparison the Deinococcus radiodurans oxidative stress tolerant catalase and superoxide dismutase genes under extracellular dGMP growth condition showed R-sq ~ 0.42 (N = 6); and the studied arsenate reductase horizontal transferred genes for toxicity survival in several microorganisms showed no correlation. Simulation results showed that R-sq < 0.4 would be improbable at less than one percent chance, suggestive of additional selection pressure when compared to the R-sq ~ 0.29 (N = 21) in the studied transferred genes in Mealybug. The mild correlation of R-sq ~ 0.5 for fractal dimension versus transcription level in the studied Deinococcus radiodurans sequences upon extracellular dGMP growth condition would suggest that lower fractal dimension with less electron density fluctuation favors higher transcription level.

  15. Targeted gene transfer into rat facial muscles by nanosecond pulsed laser-induced stress waves.

    PubMed

    Kurita, Akihiro; Matsunobu, Takeshi; Satoh, Yasushi; Ando, Takahiro; Sato, Shunichi; Obara, Minoru; Shiotani, Akihiro

    2011-09-01

    We investigate the feasibility of using nanosecond pulsed laser-induced stress waves (LISWs) for gene transfer into rat facial muscles. LISWs are generated by irradiating a black natural rubber disk placed on the target tissue with nanosecond pulsed laser light from the second harmonics (532 nm) of a Q-switched Nd:YAG laser, which is widely used in head and neck surgery and proven to be safe. After injection of plasmid deoxyribose nucleic acid (DNA) coding for Lac Z into rat facial muscles, pulsed laser is used to irradiate the laser target on the skin surface without incision or exposure of muscles. Lac Z expression is detected by X-gal staining of excised rat facial skin and muscles. Strong Lac Z expression is observed seven days after gene transfer, and sustained for up to 14 days. Gene transfer is achieved in facial muscles several millimeters deep from the surface. Gene expression is localized to the tissue exposed to LISWs. No tissue damage from LISWs is observed. LISW is a promising nonviral target gene transfer method because of its high spatial controllability, easy applicability, and minimal invasiveness. Gene transfer using LISW to produce therapeutic proteins such as growth factors could be used to treat nerve injury and paralysis. PMID:21950944

  16. Gene Transfers Shaped the Evolution of De Novo NAD+ Biosynthesis in Eukaryotes

    PubMed Central

    Ternes, Chad M.; Schönknecht, Gerald

    2014-01-01

    NAD+ is an essential molecule for life, present in each living cell. It can function as an electron carrier or cofactor in redox biochemistry and energetics, and serves as substrate to generate the secondary messenger cyclic ADP ribose and nicotinic acid adenine dinucleotide phosphate. Although de novo NAD+ biosynthesis is essential, different metabolic pathways exist in different eukaryotic clades. The kynurenine pathway starting with tryptophan was most likely present in the last common ancestor of all eukaryotes, and is active in fungi and animals. The aspartate pathway, detected in most photosynthetic eukaryotes, was probably acquired from the cyanobacterial endosymbiont that gave rise to chloroplasts. An evolutionary analysis of enzymes catalyzing de novo NAD+ biosynthesis resulted in evolutionary trees incongruent with established organismal phylogeny, indicating numerous gene transfers. Endosymbiotic gene transfers probably introduced the aspartate pathway into eukaryotes and may have distributed it among different photosynthetic clades. In addition, several horizontal gene transfers substituted eukaryotic genes with bacterial orthologs. Although horizontal gene transfer is accepted as a key mechanism in prokaryotic evolution, it is supposed to be rare in eukaryotic evolution. The essential metabolic pathway of de novo NAD+ biosynthesis in eukaryotes was shaped by numerous gene transfers. PMID:25169983

  17. Targeted gene transfer into rat facial muscles by nanosecond pulsed laser-induced stress waves

    NASA Astrophysics Data System (ADS)

    Kurita, Akihiro; Matsunobu, Takeshi; Satoh, Yasushi; Ando, Takahiro; Sato, Shunichi; Obara, Minoru; Shiotani, Akihiro

    2011-09-01

    We investigate the feasibility of using nanosecond pulsed laser-induced stress waves (LISWs) for gene transfer into rat facial muscles. LISWs are generated by irradiating a black natural rubber disk placed on the target tissue with nanosecond pulsed laser light from the second harmonics (532 nm) of a Q-switched Nd:YAG laser, which is widely used in head and neck surgery and proven to be safe. After injection of plasmid deoxyribose nucleic acid (DNA) coding for Lac Z into rat facial muscles, pulsed laser is used to irradiate the laser target on the skin surface without incision or exposure of muscles. Lac Z expression is detected by X-gal staining of excised rat facial skin and muscles. Strong Lac Z expression is observed seven days after gene transfer, and sustained for up to 14 days. Gene transfer is achieved in facial muscles several millimeters deep from the surface. Gene expression is localized to the tissue exposed to LISWs. No tissue damage from LISWs is observed. LISW is a promising nonviral target gene transfer method because of its high spatial controllability, easy applicability, and minimal invasiveness. Gene transfer using LISW to produce therapeutic proteins such as growth factors could be used to treat nerve injury and paralysis.

  18. Synthetic gene transfer vectors II: back to the future.

    PubMed

    Behr, Jean-Paul

    2012-07-17

    The discovery of RNA interference has given a new lease on life to both the chemistry of oligonucleotides and chemical approaches for the intracellular delivery of nucleic acids. In particular, delivery of siRNA, whether in vitro for screening and target validation purposes or in humans as a new class of drugs, may revolutionize our approach to therapy. Their impact could equal that of the bioproduction and various uses of monoclonal antibodies today. Unfortunately, global pharmaceutical companies again seem to be waiting to buy the next Genentech or Genzyme of gene silencing rather than investing research and development into this promising area of research. Gene silencing encounters barriers similar to gene addition and hence may benefit from the extra decade of experience brought by gene therapy. "Chemical" transfection of cells in culture has become routine, and this Account discusses some of the reasons this success has not extended to nonviral gene therapy trials, most of which do not progress beyond the phase 2 stage. The author also discusses a (much debated) mechanism of nucleic acid cell entry and subsequent release of the polycationic particles into the cytoplasm. Both topics should be useful to those interested in delivery of siRNA. The move from gene therapy toward siRNA as an oligonucleotide-based therapy strategy provides a much wider range of druggable targets. Even though these molecules are a hundredfold smaller than a gene, they are delivered via similar cellular mechanisms. Their complexes with cationic polymers are less stable than those with a higher number of phosphate groups, which may be compensated by siRNA concatemerization or by chemical conjugation with the cationic carrier. Thus chemistry is again desperately needed. PMID:22311735

  19. Optical fiber-based photomechanical gene transfer system for in vivo application.

    PubMed

    Sato, Shunichi; Ando, Takahiro; Obara, Minoru

    2011-12-01

    We developed an optical-fiber-based photomechanical gene transfer system for endoscopic or catheter-based application. A fiber tip with a laser-absorbing film covered with a transparent plastic disk for plasma confinement was attached to a quartz fiber; the film was irradiated with nanosecond laser pulses transmitted through the fiber to generate photomechanical waves (PMWs). Characteristics of PMWs emitted from the fiber tip were examined to confirm the necessary conditions for gene transfer. We then attempted to transfer reporter genes to the rat skin as a test tissue in vivo with the fiber system, and the results showed significantly high protein levels and spatially selective pinpoint gene expressions in the tissue. PMID:22139237

  20. The agricultural antibiotic carbadox induces phage-mediated gene transfer in Salmonella

    PubMed Central

    Bearson, Bradley L.; Allen, Heather K.; Brunelle, Brian W.; Lee, In Soo; Casjens, Sherwood R.; Stanton, Thaddeus B.

    2013-01-01

    Antibiotics are used for disease therapeutic or preventative effects in humans and animals, as well as for enhanced feed conversion efficiency in livestock. Antibiotics can also cause undesirable effects in microbial populations, including selection for antibiotic resistance, enhanced pathogen invasion, and stimulation of horizontal gene transfer. Carbadox is a veterinary antibiotic used in the US during the starter phase of swine production for improved feed efficiency and control of swine dysentery and bacterial swine enteritis. Carbadox has been shown in vitro to induce phage-encoded Shiga toxin in Shiga toxin-producing Escherichia coli (STEC) and a phage-like element transferring antibiotic resistance genes in Brachyspira hyodysenteriae, but the effect of carbadox on prophages in other bacteria is unknown. This study examined carbadox exposure on prophage induction and genetic transfer in Salmonella enterica serovar Typhimurium, a human foodborne pathogen that frequently colonizes swine without causing disease. S. Typhimurium LT2 exposed to carbadox induced prophage production, resulting in bacterial cell lysis and release of virions that were visible by electron microscopy. Carbadox induction of phage-mediated gene transfer was confirmed by monitoring the transduction of a sodCIII::neo cassette in the Fels-1 prophage from LT2 to a recipient Salmonella strain. Furthermore, carbadox frequently induced generalized transducing phages in multidrug-resistant phage type DT104 and DT120 isolates, resulting in the transfer of chromosomal and plasmid DNA that included antibiotic resistance genes. Our research indicates that exposure of Salmonella to carbadox induces prophages that can transfer virulence and antibiotic resistance genes to susceptible bacterial hosts. Carbadox-induced, phage-mediated gene transfer could serve as a contributing factor in bacterial evolution during animal production, with prophages being a reservoir for bacterial fitness genes in the

  1. Adenovirus-mediated gene transfer to ciliated airway epithelia requires prolonged incubation time.

    PubMed Central

    Zabner, J; Zeiher, B G; Friedman, E; Welsh, M J

    1996-01-01

    The efficiency of adenovirus-mediated gene transfer to airway epithelia will be an important factor in determining whether recombinant adenoviruses can be developed as vectors for transferring cystic fibrosis transmembrane conductance regulator (CFTR) cDNA to patients with cystic fibrosis. Current understanding of the biology of CF lung disease suggests that vectors should express transgene in mature, ciliated airway epithelia. We evaluated the efficiency of adenovirus-mediated gene transfer to primary cultures of normal and CF human airway epithelia. Our studies showed that the airway cells developed from an undifferentiated epithelium with markers characteristic of basal cells and a surface covered by short microvilli 3 days after seeding to a mature epithelium whose apical surface was covered with cilia by 10 to 14 days. The ability of adenovirus vectors to express a reporter gene and to correct defective cyclic AMP-stimulated Cl- transport in CF epithelia was correlated inversely with the state of differentiation. However, the inefficiency of adenovirus-mediated gene transfer could be partially corrected when the contact time between vector and epithelium was prolonged. After prolonged contact, we observed complete correction of the CF Cl- transport defect in differentiated CF airway epithelia in culture and of the Cl- transport defect in the nasal epithelia of mice homozygous for the deltaF508 mutation. The fact that gene transfer to airway epithelia required prolonged incubation with vector contrasts with the rapid infection observed in cell models such as 293 and HeLa cells, which are commonly used to study adenovirus infection. Gene transfer observed after prolonged incubation may result from mechanisms different from those that mediate infection of 293 cells. These observations suggest that interventions that either increase the contact time or alter the epithelium or the vector may be required to facilitate gene transfer to ciliated respiratory epithelia

  2. Prokaryotic genes in eukaryotic genome sequences: when to infer horizontal gene transfer and when to suspect an actual microbe.

    PubMed

    Artamonova, Irena I; Lappi, Tanya; Zudina, Liudmila; Mushegian, Arcady R

    2015-07-01

    Assessment of phylogenetic positions of predicted gene and protein sequences is a routine step in any genome project, useful for validating the species' taxonomic position and for evaluating hypotheses about genome evolution and function. Several recent eukaryotic genome projects have reported multiple gene sequences that were much more similar to homologues in bacteria than to any eukaryotic sequence. In the spirit of the times, horizontal gene transfer from bacteria to eukaryotes has been invoked in some of these cases. Here, we show, using comparative sequence analysis, that some of those bacteria-like genes indeed appear likely to have been horizontally transferred from bacteria to eukaryotes. In other cases, however, the evidence strongly indicates that the eukaryotic DNA sequenced in the genome project contains a sample of non-integrated DNA from the actual bacteria, possibly providing a window into the host microbiome. Recent literature suggests also that common reagents, kits and laboratory equipment may be systematically contaminated with bacterial DNA, which appears to be sampled by metagenome projects non-specifically. We review several bioinformatic criteria that help to distinguish putative horizontal gene transfers from the admixture of genes from autonomously replicating bacteria in their hosts' genome databases or from the reagent contamination. PMID:25919787

  3. Two Horizontally Transferred Xenobiotic Resistance Gene Clusters Associated with Detoxification of Benzoxazolinones by Fusarium Species

    PubMed Central

    Glenn, Anthony E.; Davis, C. Britton; Gao, Minglu; Gold, Scott E.; Mitchell, Trevor R.; Proctor, Robert H.; Stewart, Jane E.; Snook, Maurice E.

    2016-01-01

    Microbes encounter a broad spectrum of antimicrobial compounds in their environments and often possess metabolic strategies to detoxify such xenobiotics. We have previously shown that Fusarium verticillioides, a fungal pathogen of maize known for its production of fumonisin mycotoxins, possesses two unlinked loci, FDB1 and FDB2, necessary for detoxification of antimicrobial compounds produced by maize, including the γ-lactam 2-benzoxazolinone (BOA). In support of these earlier studies, microarray analysis of F. verticillioides exposed to BOA identified the induction of multiple genes at FDB1 and FDB2, indicating the loci consist of gene clusters. One of the FDB1 cluster genes encoded a protein having domain homology to the metallo-β-lactamase (MBL) superfamily. Deletion of this gene (MBL1) rendered F. verticillioides incapable of metabolizing BOA and thus unable to grow on BOA-amended media. Deletion of other FDB1 cluster genes, in particular AMD1 and DLH1, did not affect BOA degradation. Phylogenetic analyses and topology testing of the FDB1 and FDB2 cluster genes suggested two horizontal transfer events among fungi, one being transfer of FDB1 from Fusarium to Colletotrichum, and the second being transfer of the FDB2 cluster from Fusarium to Aspergillus. Together, the results suggest that plant-derived xenobiotics have exerted evolutionary pressure on these fungi, leading to horizontal transfer of genes that enhance fitness or virulence. PMID:26808652

  4. CD133-targeted gene transfer into long-term repopulating hematopoietic stem cells.

    PubMed

    Brendel, Christian; Goebel, Benjamin; Daniela, Abriss; Brugman, Martijn; Kneissl, Sabrina; Schwäble, Joachim; Kaufmann, Kerstin B; Müller-Kuller, Uta; Kunkel, Hana; Chen-Wichmann, Linping; Abel, Tobias; Serve, Hubert; Bystrykh, Leonid; Buchholz, Christian J; Grez, Manuel

    2015-01-01

    Gene therapy for hematological disorders relies on the genetic modification of CD34(+) cells, a heterogeneous cell population containing about 0.01% long-term repopulating cells. Here, we show that the lentiviral vector CD133-LV, which uses a surface marker on human primitive hematopoietic stem cells (HSCs) as entry receptor, transfers genes preferentially into cells with high engraftment capability. Transduction of unstimulated CD34(+) cells with CD133-LV resulted in gene marking of cells with competitive proliferative advantage in vitro and in immunodeficient mice. The CD133-LV-transduced population contained significantly more cells with repopulating capacity than cells transduced with vesicular stomatitis virus (VSV)-LV, a lentiviral vector pseudotyped with the vesicular stomatitis virus G protein. Upon transfer of a barcode library, CD133-LV-transduced cells sustained gene marking in vivo for a prolonged period of time with a 6.7-fold higher recovery of barcodes compared to transduced control cells. Moreover, CD133-LV-transduced cells were capable of repopulating secondary recipients. Lastly, we show that this targeting strategy can be used for transfer of a therapeutic gene into CD34(+) cells obtained from patients suffering of X-linked chronic granulomatous disease. In conclusion, direct gene transfer into CD133(+) cells allows for sustained long-term engraftment of gene corrected cells. PMID:25189742

  5. Carotenoids in unexpected places: gall midges, lateral gene transfer, and carotenoid biosynthesis in animals.

    PubMed

    Cobbs, Cassidy; Heath, Jeremy; Stireman, John O; Abbot, Patrick

    2013-08-01

    Carotenoids are conjugated isoprenoid molecules with many important physiological functions in organisms, including roles in photosynthesis, oxidative stress reduction, vision, diapause, photoperiodism, and immunity. Until recently, it was believed that only plants, microorganisms, and fungi were capable of synthesizing carotenoids and that animals acquired them from their diet, but recent studies have demonstrated that two arthropods (pea aphid and spider mite) possess a pair of genes homologous to those required for the first step of carotenoid biosynthesis. Absent in all other known animal genomes, these genes appear to have been acquired by aphids and spider mites in one or several lateral gene transfer events from a fungal donor. We report the third case of fungal carotenoid biosynthesis gene homologs in an arthropod: flies from the family Cecidomyiidae, commonly known as gall midges. Using phylogenetic analyses we show that it is unlikely that lycopene cyclase/phytoene synthase and phytoene desaturase homologs were transferred singly to an ancient arthropod ancestor; instead we propose that genes were transferred independently from related fungal donors after divergence of the major arthropod lineages. We also examine variation in intron placement and copy number of the carotenoid genes that may underlie function in the midges. This trans-kingdom transfer of carotenoid genes may represent a key innovation, underlying the evolution of phytophagy and plant-galling in gall midges and facilitating their extensive diversification across plant lineages. PMID:23542649

  6. Two Horizontally Transferred Xenobiotic Resistance Gene Clusters Associated with Detoxification of Benzoxazolinones by Fusarium Species.

    PubMed

    Glenn, Anthony E; Davis, C Britton; Gao, Minglu; Gold, Scott E; Mitchell, Trevor R; Proctor, Robert H; Stewart, Jane E; Snook, Maurice E

    2016-01-01

    Microbes encounter a broad spectrum of antimicrobial compounds in their environments and often possess metabolic strategies to detoxify such xenobiotics. We have previously shown that Fusarium verticillioides, a fungal pathogen of maize known for its production of fumonisin mycotoxins, possesses two unlinked loci, FDB1 and FDB2, necessary for detoxification of antimicrobial compounds produced by maize, including the γ-lactam 2-benzoxazolinone (BOA). In support of these earlier studies, microarray analysis of F. verticillioides exposed to BOA identified the induction of multiple genes at FDB1 and FDB2, indicating the loci consist of gene clusters. One of the FDB1 cluster genes encoded a protein having domain homology to the metallo-β-lactamase (MBL) superfamily. Deletion of this gene (MBL1) rendered F. verticillioides incapable of metabolizing BOA and thus unable to grow on BOA-amended media. Deletion of other FDB1 cluster genes, in particular AMD1 and DLH1, did not affect BOA degradation. Phylogenetic analyses and topology testing of the FDB1 and FDB2 cluster genes suggested two horizontal transfer events among fungi, one being transfer of FDB1 from Fusarium to Colletotrichum, and the second being transfer of the FDB2 cluster from Fusarium to Aspergillus. Together, the results suggest that plant-derived xenobiotics have exerted evolutionary pressure on these fungi, leading to horizontal transfer of genes that enhance fitness or virulence. PMID:26808652

  7. Guanidinylated block copolymers for gene transfer: A comparison with amine-based materials for in vitro and in vivo gene transfer efficiency

    PubMed Central

    Choi, Jennifer L.; Tan, James-Kevin Y.; Sellers, Drew L.; Wei, Hua; Horner, Philip J.; Pun, Suzie H.

    2015-01-01

    There is currently no cure for neuron loss in the brain, which can occur due to traumatic injury or neurodegenerative disease. One method proposed to enhance neurogenesis in the brain is gene transfer to neural progenitor cells. In this work, a guanidine-based copolymer was synthesized and compared to an amine-based copolymer analog previously shown to effectively deliver genes in the murine brain. The guanidine-based copolymer was more efficient at gene transfer to immortalized, cultured cell lines; however, the amine-based copolymer was more effective at gene transfer in the brain. DNA condensation studies revealed that the nucleic acid complexes formed with the guanidine-based copolymer were more susceptible to unpackaging in the presence of heparin sulfate proteoglycans compared to complexes formed with the amine-based copolymer. Therefore, polyplexes formed from the amine-based copolymer may be more resistant to destabilization by the heparan sulfate proteoglycans present in the stem cell niches of the brain. PMID:25907042

  8. Horizontal transfer of archaeal genes into the deinococcaceae: detection by molecular and computer-based approaches

    NASA Technical Reports Server (NTRS)

    Olendzenski, L.; Liu, L.; Zhaxybayeva, O.; Murphey, R.; Shin, D. G.; Gogarten, J. P.

    2000-01-01

    Members of the Deinococcaceae (e.g., Thermus, Meiothermus, Deinococcus) contain A/V-ATPases typically found in Archaea or Eukaryotes which were probably acquired by horizontal gene transfer. Two methods were used to quantify the extent to which archaeal or eukaryotic genes have been acquired by this lineage. Screening of a Meiothermus ruber library with probes made against Thermoplasma acidophilum DNA yielded a number of clones which hybridized more strongly than background. One of these contained the prolyl tRNA synthetase (RS) gene. Phylogenetic analysis shows the M. ruber and D. radiodurans prolyl RS to be more closely related to archaeal and eukaryal forms of this gene than to the typical bacterial type. Using a bioinformatics approach, putative open reading frames (ORFs) from the prerelease version of the D. radiodurans genome were screened for genes more closely related to archaeal or eukaryotic genes. Putative ORFs were searched against representative genomes from each of the three domains using automated BLAST. ORFs showing the highest matches against archaeal and eukaryotic genes were collected and ranked. Among the top-ranked hits were the A/V-ATPase catalytic and noncatalytic subunits and the prolyl RS genes. Using phylogenetic methods, ORFs were analyzed and trees assessed for evidence of horizontal gene transfer. Of the 45 genes examined, 20 showed topologies in which D. radiodurans homologues clearly group with eukaryotic or archaeal homologues, and 17 additional trees were found to show probable evidence of horizontal gene transfer. Compared to the total number of ORFs in the genome, those that can be identified as having been acquired from Archaea or Eukaryotes are relatively few (approximately 1%), suggesting that interdomain transfer is rare.

  9. Horizontal Gene Transfer and the Genomics of Enterococcal Antibiotic Resistance

    PubMed Central

    Palmer, Kelli L.; Kos, Veronica N.

    2010-01-01

    Summary Enterococci are Gram-positive bacteria that normally colonize gastrointestinal tracts of humans and animals. They are of growing concern because of their ability to cause antibiotic resistant hospital infections. Antibiotic resistance has been acquired, and has disseminated throughout enterococci, via horizontal transfer of mobile genetic elements. This transmission has been mediated mainly by conjugative plasmids of the pheromone-responsive and broad host range incompatibility group 18 type. Genome sequencing is revealing the extent of diversity of these and other mobile elements in enterococci, as well as the extent of recombination and rearrangement resulting in new phenotypes. Pheromone-responsive plasmids were recently shown to promote genome plasticity in antibiotic resistant Enterococcus faecalis, and their involvement has been implicated in E. faecium as well. Further, incompatibility group 18 plasmids have recently played an important role in mediating transfer of vancomycin resistance from enterococci to methicillin resistant strains of S. aureus. PMID:20837397

  10. Gene Transfer to the Desiccation-Tolerant Cyanobacterium Chroococcidiopsis

    PubMed Central

    Billi, Daniela; Friedmann, E. Imre; Helm, Richard F.; Potts, Malcolm

    2001-01-01

    The coccoid cyanobacterium Chroococcidiopsis dominates microbial communities in the most extreme arid hot and cold deserts. These communities withstand constraints that result from multiple cycles of drying and wetting and/or prolonged desiccation, through mechanisms which remain poorly understood. Here we describe the first system for genetic manipulation of Chroococcidiopsis. Plasmids pDUCA7 and pRL489, based on the pDU1 replicon of Nostoc sp. strain PCC 7524, were transferred to different isolates of Chroococcidiopsis via conjugation and electroporation. This report provides the first evidence that pDU1 replicons can be maintained in cyanobacteria other than Nostoc and Anabaena. Following conjugation, both plasmids replicated in Chroococcidiopsis sp. strains 029, 057, and 123 but not in strains 171 and 584. Both plasmids were electroporated into strains 029 and 123 but not into strains 057, 171, and 584. Expression of PpsbA-luxAB on pRL489 was visualized through in vivo luminescence. Efficiencies of conjugative transfer for pDUCA7 and pRL489 into Chroococcidiopsis sp. strain 029 were approximately 10−2 and 10−4 transconjugants per recipient cell, respectively. Conjugative transfer occurred with a lower efficiency into strains 057 and 123. Electrotransformation efficiencies of about 10−4 electrotransformants per recipient cell were achieved with strains 029 and 123, using either pDUCA7 or pRL489. Extracellular deoxyribonucleases were associated with each of the five strains. Phylogenetic analysis, based upon the V6 to V8 variable regions of 16S rRNA, suggests that desert strains 057, 123, 171, and 029 are distinct from the type species strain Chroococcidiopsis thermalis PCC 7203. The high efficiency of conjugative transfer of Chroococcidiopsis sp. strain 029, from the Negev Desert, Israel, makes this a suitable experimental strain for genetic studies on desiccation tolerance. PMID:11244070

  11. Micro RNA-98 interferes with expression interleukin-10 in peripheral B cells of patients with lung cancer

    PubMed Central

    Li, Yun; Rong, Jian; Qin, Jie; He, Jin-yuan; Chen, Hui-guo; Huang, Shao-hong

    2016-01-01

    Interleukin (IL)-10-producing B cells (B10 cells) plays an important role in the tumor tolerance. High frequency of peripheral B10 cell was reported in patients with lung cancer recently. Micro RNA (miR) regulates some gene expression. This study test a hypothesis that miR-98 suppresses the expression of IL-10 in B cells of subjects with lung cancer. The results showed that the levels of miR-98 were significantly less in peripheral B cells of patients with lung cancer than that in healthy subjects. IL-10 mRNA levels in peripheral B cells were significantly higher in lung cancer patients as compared with healthy controls. A negative correlation was identified between miR-98 and IL-10 in peripheral B cells. Serum IL-13 was higher in lung cancer patients than that in healthy controls. The levels of IL-13 were also negatively correlated with IL-10 in B cells. Exposure B10 cells to IL-13 in the culture or over expression of miR-98 reduced the expression of IL-10 in B cells. Administration with miR-98-laden liposomes inhibited the lung cancer growth in a mouse model. In conclusion, up regulation of miR-98 inhibits the expression of IL-10 in B cells, which may contribute to inhibit the lung cancer tolerance in the body. PMID:27605397

  12. Micro RNA-98 interferes with expression interleukin-10 in peripheral B cells of patients with lung cancer.

    PubMed

    Li, Yun; Rong, Jian; Qin, Jie; He, Jin-Yuan; Chen, Hui-Guo; Huang, Shao-Hong

    2016-01-01

    Interleukin (IL)-10-producing B cells (B10 cells) plays an important role in the tumor tolerance. High frequency of peripheral B10 cell was reported in patients with lung cancer recently. Micro RNA (miR) regulates some gene expression. This study test a hypothesis that miR-98 suppresses the expression of IL-10 in B cells of subjects with lung cancer. The results showed that the levels of miR-98 were significantly less in peripheral B cells of patients with lung cancer than that in healthy subjects. IL-10 mRNA levels in peripheral B cells were significantly higher in lung cancer patients as compared with healthy controls. A negative correlation was identified between miR-98 and IL-10 in peripheral B cells. Serum IL-13 was higher in lung cancer patients than that in healthy controls. The levels of IL-13 were also negatively correlated with IL-10 in B cells. Exposure B10 cells to IL-13 in the culture or over expression of miR-98 reduced the expression of IL-10 in B cells. Administration with miR-98-laden liposomes inhibited the lung cancer growth in a mouse model. In conclusion, up regulation of miR-98 inhibits the expression of IL-10 in B cells, which may contribute to inhibit the lung cancer tolerance in the body. PMID:27605397

  13. Transferring Sclerotinia Resistance Genes from Wild Helianthus into Cultivated Sunflower

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To enhance resistance to Sclerotinia head and stalk rot in cultivated sunflower, mining and introgression of Sclerotinia resistance genes from diverse wild Helianthus accessions into cultivated sunflower has been conducted using backcrossing method since 2004. During the last four years, numerous in...

  14. Massive Gene Transfer and Extensive RNA Editing of a Symbiotic Dinoflagellate Plastid Genome

    PubMed Central

    Mungpakdee, Sutada; Shinzato, Chuya; Takeuchi, Takeshi; Kawashima, Takeshi; Koyanagi, Ryo; Hisata, Kanako; Tanaka, Makiko; Goto, Hiroki; Fujie, Manabu; Lin, Senjie; Satoh, Nori; Shoguchi, Eiichi

    2014-01-01

    Genome sequencing of Symbiodinium minutum revealed that 95 of 109 plastid-associated genes have been transferred to the nuclear genome and subsequently expanded by gene duplication. Only 14 genes remain in plastids and occur as DNA minicircles. Each minicircle (1.8–3.3 kb) contains one gene and a conserved noncoding region containing putative promoters and RNA-binding sites. Nine types of RNA editing, including a novel G/U type, were discovered in minicircle transcripts but not in genes transferred to the nucleus. In contrast to DNA editing sites in dinoflagellate mitochondria, which tend to be highly conserved across all taxa, editing sites employed in DNA minicircles are highly variable from species to species. Editing is crucial for core photosystem protein function. It restores evolutionarily conserved amino acids and increases peptidyl hydropathy. It also increases protein plasticity necessary to initiate photosystem complex assembly. PMID:24881086

  15. Mucus altering agents as adjuncts for nonviral gene transfer to airway epithelium.

    PubMed

    Ferrari, S; Kitson, C; Farley, R; Steel, R; Marriott, C; Parkins, D A; Scarpa, M; Wainwright, B; Evans, M J; Colledge, W H; Geddes, D M; Alton, E W

    2001-09-01

    Nonviral vectors have been shown to be a safe and valid alternative to recombinant viruses for gene therapy of cystic fibrosis (CF). Nevertheless, gene transfer efficiency needs to be increased before clinical efficacy is likely in man. One barrier to increased efficacy is normal airway mucus. Using an ex vivo model of sheep tracheal epithelium, we show that this barrier can, in part, be overcome by treatment with the mucolytic agents, Nacystelyn or N-acetylcysteine using either a cationic lipid or a cationic polymer as the gene transfer agent. Further, in vivo application of either Nacystelyn or the anticholinergic glycopyrrolate, both clinically used agents, resulted in increased reporter gene expression in the mouse lung, but no significant correction of the bioelectric defect in CF null mice. These results, whilst unlikely to be sufficient in themselves to achieve clinically relevant gene therapy, may be a further useful step in the attainment of this goal. PMID:11571577

  16. HGT-Finder: A New Tool for Horizontal Gene Transfer Finding and Application to Aspergillus genomes

    PubMed Central

    Nguyen, Marcus; Ekstrom, Alex; Li, Xueqiong; Yin, Yanbin

    2015-01-01

    Horizontal gene transfer (HGT) is a fast-track mechanism that allows genetically unrelated organisms to exchange genes for rapid environmental adaptation. We developed a new phyletic distribution-based software, HGT-Finder, which implements a novel bioinformatics algorithm to calculate a horizontal transfer index and a probability value for each query gene. Applying this new tool to the Aspergillus fumigatus, Aspergillus flavus, and Aspergillus nidulans genomes, we found 273, 542, and 715 transferred genes (HTGs), respectively. HTGs have shorter length, higher guanine-cytosine (GC) content, and relaxed selection pressure. Metabolic process and secondary metabolism functions are significantly enriched in HTGs. Gene clustering analysis showed that 61%, 41% and 74% of HTGs in the three genomes form physically linked gene clusters (HTGCs). Overlapping manually curated, secondary metabolite gene clusters (SMGCs) with HTGCs found that 9 of the 33 A. fumigatus SMGCs and 31 of the 65 A. nidulans SMGCs share genes with HTGCs, and that HTGs are significantly enriched in SMGCs. Our genome-wide analysis thus presented very strong evidence to support the hypothesis that HGT has played a very critical role in the evolution of SMGCs. The program is freely available at http://cys.bios.niu.edu/HGTFinder/HGTFinder.tar.gz. PMID:26473921

  17. GFP as a marker for transient gene transfer and expression in Mycoplasma hyorhinis.

    PubMed

    Ishag, Hassan Z A; Liu, Maojun; Yang, Ruosong; Xiong, Qiyan; Feng, Zhixin; Shao, Guoqing

    2016-01-01

    Mycoplasma hyorhinis (M. hyorhinis) is an opportunistic pathogen of pigs and has been shown to transform cell cultures, which has increased the interest of researchers. The green florescence proteins (GFP) gene of Aquorea victoria, proved to be a vital marker to identify transformed cells in mixed populations. Use of GFP to observe gene transfer and expression in M. hyorhinis (strain HUB-1) has not been described. We have constructed a pMD18-O/MHRgfp plasmid containing the p97 gene promoter, origin of replication, tetracycline resistance marker and GFP gene controlled by the p97 gene promoter. The plasmid transformed into M. hyorhinis with a frequency of ~4 × 10(-3) cfu/µg plasmid DNA and could be detected by PCR amplification of the GFP gene from the total DNA of the transformant mycoplasmas. Analysis of a single clone grown on KM2-Agar containing tetracycline, showed a green fluorescence color. Conclusively, this report suggests the usefulness of GFP to monitor transient gene transfer and expression in M. hyorhinis, eventually minimizing screening procedures for gene transfer and expression. PMID:27386255

  18. Plant expansins in bacteria and fungi: evolution by horizontal gene transfer and independent domain fusion.

    PubMed

    Nikolaidis, Nikolas; Doran, Nicole; Cosgrove, Daniel J

    2014-02-01

    Horizontal gene transfer (HGT) has been described as a common mechanism of transferring genetic material between prokaryotes, whereas genetic transfers from eukaryotes to prokaryotes have been rarely documented. Here we report a rare case of HGT in which plant expansin genes that code for plant cell-wall loosening proteins were transferred from plants to bacteria, fungi, and amoebozoa. In several cases, the species in which the expansin gene was found is either in intimate association with plants or is a known plant pathogen. Our analyses suggest that at least two independent genetic transfers occurred from plants to bacteria and fungi. These events were followed by multiple HGT events within bacteria and fungi. We have also observed that in bacteria expansin genes have been independently fused to DNA fragments that code for an endoglucanase domain or for a carbohydrate binding module, pointing to functional convergence at the molecular level. Furthermore, the functional similarities between microbial expansins and their plant xenologs suggest that these proteins mediate microbial-plant interactions by altering the plant cell wall and therefore may provide adaptive advantages to these species. The evolution of these nonplant expansins represents a unique case in which bacteria and fungi have found innovative and adaptive ways to interact with and infect plants by acquiring genes from their host. This evolutionary paradigm suggests that despite their low frequency such HGT events may have significantly contributed to the evolution of prokaryotic and eukaryotic species. PMID:24150040

  19. Interleukin 10 and dendritic cells are the main suppression mediators of regulatory T cells in human neurocysticercosis.

    PubMed

    Arce-Sillas, A; Álvarez-Luquín, D D; Cárdenas, G; Casanova-Hernández, D; Fragoso, G; Hernández, M; Proaño Narváez, J V; García-Vázquez, F; Fleury, A; Sciutto, E; Adalid-Peralta, L

    2016-02-01

    Neurocysticercosis is caused by the establishment of Taenia solium cysticerci in the central nervous system. It is considered that, during co-evolution, the parasite developed strategies to modulate the host's immune response. The action mechanisms of regulatory T cells in controlling the immune response in neurocysticercosis are studied in this work. Higher blood levels of regulatory T cells with CD4(+) CD45RO(+) forkhead box protein 3 (FoxP3)(high) and CD4(+) CD25(high) FoxP3(+) CD95(high) phenotype and of non-regulatory CD4(+) CD45RO(+) FoxP3(med) T cells were found in neurocysticercosis patients with respect to controls. Interestingly, regulatory T cells express higher levels of cytotoxic T lymphocyte antigen 4 (CTLA-4), lymphocyte-activation gene 3 (LAG-3), programmed death 1 (PD-1) and glucocorticoid-induced tumour necrosis factor receptor (GITR), suggesting a cell-to-cell contact mechanism with dendritic cells. Furthermore, higher IL-10 and regulatory T cell type 1 (Tr1) levels were found in neurocysticercosis patients' peripheral blood, suggesting that the action mechanism of regulatory T cells involves the release of immunomodulatory cytokines. No evidence was found of the regulatory T cell role in inhibiting the proliferative response. Suppressive regulatory T cells from neurocysticercosis patients correlated negatively with late activated lymphocytes (CD4(+) CD38(+) ). Our results suggest that, during neurocysticercosis, regulatory T cells could control the immune response, probably by a cell-to-cell contact with dendritic cells and interleukin (IL)-10 release by Tr1, to create an immunomodulatory environment that may favour the development of T. solium cysticerci and their permanence in the central nervous system. PMID:26391104

  20. Exploration of new perspectives and limitations in Agrobacterium mediated gene transfer technology. Progress report, [June 1, 1992-- May 31, 1994

    SciTech Connect

    Marton, L.

    1994-12-31

    This report describes progress aimed at constructing gene-transfer technology for Nicotiana plumbaginifolia. Most actual effort as described herein has so far been directed at exploring new perspectives and limitations in Agrobacterium mediated gene transfer. Accomplishments are described using a core homologous gene targeting vector.

  1. Rare Events of Intragenus and Intraspecies Horizontal Transfer of the 16S rRNA Gene

    PubMed Central

    Tian, Ren-Mao; Cai, Lin; Zhang, Wei-Peng; Cao, Hui-Luo; Qian, Pei-Yuan

    2015-01-01

    Horizontal gene transfer (HGT) of operational genes has been widely reported in prokaryotic organisms. However, informational genes such as those involved in transcription and translation processes are very difficult to be horizontally transferred, as described by Woese’s complexity hypothesis. Here, we analyzed all of the completed prokaryotic genome sequences (2,143 genomes) in the NCBI (National Center for Biotechnology Information) database, scanned for genomes with high intragenomic heterogeneity of 16S rRNA gene copies, and explored potential HGT events of ribosomal RNA genes based on the phylogeny, genomic organization, and secondary structures of the ribosomal RNA genes. Our results revealed 28 genomes with relatively high intragenomic heterogeneity of multiple 16S rRNA gene copies (lowest pairwise identity <98.0%), and further analysis revealed HGT events and potential donors of the heterogeneous copies (such as HGT from Chlamydia suis to Chlamydia trachomatis) and mutation events of some heterogeneous copies (such as Streptococcus suis JS14). Interestingly, HGT of the 16S rRNA gene only occurred at intragenus or intraspecies levels, which is quite different from the HGT of operational genes. Our results improve our understanding regarding the exchange of informational genes. PMID:26220935

  2. Direct phylogenetic evidence for lateral transfer of elongation factor-like gene

    PubMed Central

    Kamikawa, Ryoma; Inagaki, Yuji; Sako, Yoshihiko

    2008-01-01

    Genes encoding elongation factor-like (EFL) proteins, which show high similarity to elongation factor-1α (EF-1α), have been found in phylogenetically distantly related eukaryotes. The sporadic distribution of “EFL-containing” lineages within “EF-1α-containing” lineages indirectly, but strongly, suggests lateral gene transfer as the principal driving force in EFL evolution. However, one of the most critical aspects in the above hypothesis, the donor lineages in any putative cases of lateral EFL gene transfer, remained unclear. In this study, we provide direct evidence for lateral transfer of an EFL gene through the analyses of 10 diatom EFL genes. All diatom EFL homologues tightly clustered in phylogenetic analyses, suggesting acquisition of the exogenous EFL gene early in diatom evolution. Our survey additionally identified Thalassiosira pseudonana as a eukaryote bearing EF-1α and EFL genes and secondary EFL gene loss in Phaeodactylum tricornutum, the complete genome of which encodes only the EF-1α gene. Most importantly, the EFL phylogeny recovered a robust grouping of homologues from diatoms, the cercozoan Bigelowiella natans, and the foraminifer Planoglabratella opecularis, with the diatoms nested within the Bigelowiella plus Planoglabratella (Rhizaria) grouping. The particular relationships recovered are further consistent with two characteristic sequence motifs. The best explanation of our data analyses is an EFL gene transfer from a foraminifer to a diatom, the first case in which the donor–recipient relationship was clarified. Finally, based on a reverse transcriptase quantitative PCR assay and the genome information of Thalassiosira and Phaeodactylum, we propose the loss of elongation factor function in Thalassiosira EF-1α. PMID:18458344

  3. Improving lipoprotein profiles by liver-directed gene transfer of low density lipoprotein receptor gene in hypercholesterolaemia mice.

    PubMed

    Ou, Hailong; Zhang, Qinghai; Zeng, Jia

    2016-06-01

    The defect of low density lipoprotein receptor disturbs cholesterol metabolism and causes familial hypercholesterolaemia (FH). In this study, we directly delivered exogenous Ldlr gene into the liver of FH model mice (Ldlr(-/-)) by lentiviral gene transfer system. The results showed that the Ldlr gene controlled by hepatocyte-specific human thyroxine-binding globulin (TBG) promoter successfully and exclusively expressed in livers.We found that, although, the content of high density lipoprotein in serum was not significantly affected by the Ldlr gene expression, the serum low density lipoprotein level was reduced by 46%, associated with a 30% and 28% decrease in triglyceride and total cholesterol, respectively, compared to uninjected Ldlr(-/-) mice. Moreover, the TBG directed expression of Ldlr significantly decreased the lipid accumulation in liver and reduced plaque burden in aorta (32%). Our results indicated that the hepatocyte-specific expression of Ldlr gene strikingly lowered serum lipid levels and resulted in amelioration of hypercholesterolaemia. PMID:27350674

  4. The transfer and transformation of collective network information in gene-matched networks

    PubMed Central

    Kitsukawa, Takashi; Yagi, Takeshi

    2015-01-01

    Networks, such as the human society network, social and professional networks, and biological system networks, contain vast amounts of information. Information signals in networks are distributed over nodes and transmitted through intricately wired links, making the transfer and transformation of such information difficult to follow. Here we introduce a novel method for describing network information and its transfer using a model network, the Gene-matched network (GMN), in which nodes (neurons) possess attributes (genes). In the GMN, nodes are connected according to their expression of common genes. Because neurons have multiple genes, the GMN is cluster-rich. We show that, in the GMN, information transfer and transformation were controlled systematically, according to the activity level of the network. Furthermore, information transfer and transformation could be traced numerically with a vector using genes expressed in the activated neurons, the active-gene array, which was used to assess the relative activity among overlapping neuronal groups. Interestingly, this coding style closely resembles the cell-assembly neural coding theory. The method introduced here could be applied to many real-world networks, since many systems, including human society and various biological systems, can be represented as a network of this type. PMID:26450411

  5. Eukaryote to gut bacteria transfer of a glycoside hydrolase gene essential for starch breakdown in plants

    PubMed Central

    Arias, Maria Cecilia; Danchin, Étienne G.J.; Coutinho, Pedro; Henrissat, Bernard; Ball, Steven

    2012-01-01

    Lateral gene transfer (LGT) between bacteria constitutes a strong force in prokaryote evolution, transforming the hierarchical tree of life into a network of relationships between species. In contrast, only a few cases of LGT from eukaryotes to prokaryotes have been reported so far. The distal animal intestine is predominantly a bacterial ecosystem, supplying the host with energy from dietary polysaccharides through carbohydrate-active enzymes absent from its genome. It has been suggested that LGT is particularly important for the human microbiota evolution. Here we show evidence for the first eukaryotic gene identified in multiple gut bacterial genomes. We found in the genome sequence of several gut bacteria, a typically eukaryotic glycoside-hydrolase necessary for starch breakdown in plants. The distribution of this gene is patchy in gut bacteria with presence otherwise detected only in a few environmental bacteria. We speculate that the transfer of this gene to gut bacteria occurred by a sequence of two key LGT events; first, an original eukaryotic gene was transferred probably from Archaeplastida to environmental bacteria specialized in plant polysaccharides degradation and second, the gene was transferred from the environmental bacteria to gut microbes. PMID:22934241

  6. The transfer and transformation of collective network information in gene-matched networks.

    PubMed

    Kitsukawa, Takashi; Yagi, Takeshi

    2015-01-01

    Networks, such as the human society network, social and professional networks, and biological system networks, contain vast amounts of information. Information signals in networks are distributed over nodes and transmitted through intricately wired links, making the transfer and transformation of such information difficult to follow. Here we introduce a novel method for describing network information and its transfer using a model network, the Gene-matched network (GMN), in which nodes (neurons) possess attributes (genes). In the GMN, nodes are connected according to their expression of common genes. Because neurons have multiple genes, the GMN is cluster-rich. We show that, in the GMN, information transfer and transformation were controlled systematically, according to the activity level of the network. Furthermore, information transfer and transformation could be traced numerically with a vector using genes expressed in the activated neurons, the active-gene array, which was used to assess the relative activity among overlapping neuronal groups. Interestingly, this coding style closely resembles the cell-assembly neural coding theory. The method introduced here could be applied to many real-world networks, since many systems, including human society and various biological systems, can be represented as a network of this type. PMID:26450411

  7. Mathematical modelling of antimicrobial resistance in agricultural waste highlights importance of gene transfer rate.

    PubMed

    Baker, Michelle; Hobman, Jon L; Dodd, Christine E R; Ramsden, Stephen J; Stekel, Dov J

    2016-04-01

    Antimicrobial resistance is of global concern. Most antimicrobial use is in agriculture; manures and slurry are especially important because they contain a mix of bacteria, including potential pathogens, antimicrobial resistance genes and antimicrobials. In many countries, manures and slurry are stored, especially over winter, before spreading onto fields as organic fertilizer. Thus, these are a potential location for gene exchange and selection for resistance. We develop and analyse a mathematical model to quantify the spread of antimicrobial resistance in stored agricultural waste. We use parameters from a slurry tank on a UK dairy farm as an exemplar. We show that the spread of resistance depends in a subtle way on the rates of gene transfer and antibiotic inflow. If the gene transfer rate is high, then its reduction controls resistance, while cutting antibiotic inflow has little impact. If the gene transfer rate is low, then reducing antibiotic inflow controls resistance. Reducing length of storage can also control spread of resistance. Bacterial growth rate, fitness costs of carrying antimicrobial resistance and proportion of resistant bacteria in animal faeces have little impact on spread of resistance. Therefore, effective treatment strategies depend critically on knowledge of gene transfer rates. PMID:26906100

  8. Gene gun transferring-bone morphogenetic protein 2 (BMP-2) gene enhanced bone fracture healing in rabbits

    PubMed Central

    Li, Wenju; Wei, Haifeng; Xia, Chunmei; Zhu, Xiaomeng; Hou, Guozhu; Xu, Feng; Song, Xinghua; Zhan, Yulin

    2015-01-01

    Purpose: Transferring the bone morphogenetic protein 2 (BMP-2) genes into the tissues or cells can improve the bone healing of the fracture has been widely accepted. We evaluated the efficiency of using gene gun to transfer the BMP-2 gene thereby affected the healing of a fractured bone. Methods: The vector coding for BMP-2 was constructed by a non-replicating encephalo-myocarditis virus (ECMV)-based vector. The segmental bone defect (1.5 cm) model was created by a wire-saw at the middle part of the radius bone of the New Zealand white rabbits. Then either BMP-2 gene or control vector without BMP-2 gene was injected into the tissues around the fracture site. Healing of the defects was monitored radiographically for 9 weeks, bone consolidation was determined by the Lane-Sandhu score pre- and post-operatively, which can evaluated bone formation, bone connect and bone plasticity. Results: The radiographic score and bone consolidation rates were significantly higher in animals injected with BMP-2 gene group as compared with control vector-injected animals (P<0.05). The control group still showed no radiological signs of stable healing. Western-blot and RT-PCR showed BMP-2 expression was significant increase in the tissues around the site of osseous lesions in comparison with the control vector-injected animals (P<0.05). Conclusions: Our results suggested that BMP-2 gene transferred by gene gun could increase the expression of BMP-2 protein and improved the bone callus formation therefore shortened the time of bone defect healing. PMID:26884910

  9. Adenovirus-mediated gene transfer to tumor cells.

    PubMed

    Cascalló, Manel; Alemany, Ramon

    2004-01-01

    Cell transduction in vitro is only the first step toward proving that a genetherapy vector can be useful to treat tumors. However, tumor targeting in vivo is now the milestone for gene therapy to succeed against disseminated cancer. Therefore, most valuable information is obtained from studies of vector biodistribution. Owing to the hepatotropism of adenoviral vectors, a particularly important parameter is the tumor/liver ratio. This ratio can be given at the level of gene expression if the amount of transgene expression is measured. To optimize the targeting, however, the levels of viral particles that reach the tumor compared to other organs must be studied. Most of this chapter deals with methods to quantify the virus fate in tumor-bearing animals. We present a radioactive labeling method that can be used to study biodistribution. After a small section dealing with tumor models, we describe methods to quantify different parameters related to adenovirus-mediated tumor targeting. PMID:14970588

  10. Association of Interleukin-6 and Interleukin-10 Genotypes With Radiographic Damage in Rheumatoid Arthritis Is Dependent on Autoantibody Status

    PubMed Central

    Marinou, I; Healy, J; Mewar, D; Moore, D J; Dickson, M C; Binks, M H; Montgomery, D S; Walters, K; Wilson, A G

    2007-01-01

    Objective Recent evidence has highlighted a major genetic contribution to radiographic damage in rheumatoid arthritis (RA). The objective of this study was to determine whether genetic variants in the loci for interleukin-1 (IL-1), IL-6, IL-10, protein tyrosine phosphatase N22 (PTPN22), and selenoprotein S are associated with radiographic damage. Methods Modified Larsen scores of radiographic damage were determined in a cross-sectional population of patients with RA (n = 964). Rheumatoid factor (RF) and anti–cyclic citrullinated peptide (anti-CCP) were also assayed. The Kruskal-Wallis nonparametric test was used to compare median radiographic damage scores across genotype groups, followed by the Cuzick nonparametric test for trend to assess gene-dose effects. Results An allele-dose association of IL-6 −174G with increasing radiographic damage was present (P = 0.005), but only in patients who were RF positive (P = 0.004) or anti-CCP positive (P = 0.01). Patients with the IL-10 −592CC genotype had more extensive radiographic damage than did those with the AC or AA genotype (P = 0.006), but this was observed only among patients who were RF negative (P = 0.002) or anti-CCP negative (P = 0.002). However, RF status and anti-CCP status were not associated with the IL-6 or IL-10 genotype. No other genetic associations were detected, apart from a marginal association of PTPN22 +1858T with increased radiographic damage. Conclusion The reported associations of IL-6 −174G with high IL-6 production and IL-10 −592 with low IL-10 production and our own results support a role of genetically determined dysregulated cytokine production in disease severity. The lack of association of these genotypes with RF and anti-CCP antibody status suggests that they act downstream of autoantibody production. We conclude that IL-6 and IL-10 genotypes may be useful in predicting disease severity in autoantibody-positive and autoantibody-negative patients, respectively. PMID:17665434

  11. Decreasing the effects of horizontal gene transfer on bacterial phylogeny: the Escherichia coli case study.

    PubMed

    Escobar-Páramo, Patricia; Sabbagh, Audrey; Darlu, Pierre; Pradillon, Olivier; Vaury, Christelle; Denamur, Erick; Lecointre, Guillaume

    2004-01-01

    Phylogenetic reconstructions of bacterial species from DNA sequences are hampered by the existence of horizontal gene transfer. One possible way to overcome the confounding influence of such movement of genes is to identify and remove sequences which are responsible for significant character incongruence when compared to a reference dataset free of horizontal transfer (e.g., multilocus enzyme electrophoresis, restriction fragment length polymorphism, or random amplified polymorphic DNA) using the incongruence length difference (ILD) test of Farris et al. [Cladistics 10 (1995) 315]. As obtaining this "whole genome dataset" prior to the reconstruction of a phylogeny is clearly troublesome, we have tested alternative approaches allowing the release from such reference dataset, designed for a species with modest level of horizontal gene transfer, i.e., Escherichia coli. Eleven different genes available or sequenced in this work were studied in a set of 30 E. coli reference (ECOR) strains. Either using ILD to test incongruence between each gene against the all remaining (in this case 10) genes in order to remove sequences responsible for significant incongruence, or using just a simultaneous analysis without removals, gave robust phylogenies with slight topological differences. The use of the ILD test remains a suitable method for estimating the level of horizontal gene transfer in bacterial species. Supertrees also had suitable properties to extract the phylogeny of strains, because the way they summarize taxonomic congruence clearly limits the impact of individual gene transfers on the global topology. Furthermore, this work allowed a significant improvement of the accuracy of the phylogeny within E. coli. PMID:15022774

  12. Microbubble-Enhanced Ultrasound Gene Transfer into Fibroblast Cells

    NASA Astrophysics Data System (ADS)

    Hirayama, Kota; Kaneko, Yukio; Tei, Yuichi; Matsumoto, Yoichiro

    2007-05-01

    Ultrasound finds many applications in the medical field, including ultrasound imaging, non-invasive treatment of tumors and lithotripsy. Ultrasound also has a potential to deliver some therapeutic materials, such as genes, drugs or proteins into cells. It is known that microbubbles can improve the delivery efficiency. It is believed that therapeutic materials can pass through the cell membrane whose permeability is increased by microbubble destruction or the ultrasound pressure. In this study, we investigated the delivery of GFP plasmid gene into the fibroblast cells. Ultrasound (frequency = 2.1 MHz, duty cycle = 10%) was used to irradiate the cultured cells through a medium that contains microbubbles and GFP plasmid. GFP plasmid transfection could be easily observed by fluorescence microscopy. Ultrasound irradiation under a variety of conditions resulted in successful GFP plasmid delivery. Microbubbles enhanced GFP transfection, and conclusions were drawn as to the relationship between gene transfection and various ultrasound exposure parameters. We also investigated the effect of ultrasound intensity on cell viability.

  13. Gene Transfer into Rat Brain Using Adenoviral Vectors

    PubMed Central

    Puntel, Mariana; Kroeger, Kurt M.; Sanderson, Nicholas S.R.; Thomas, Clare E.; Castro, Maria G.; Lowenstein, Pedro R.

    2010-01-01

    Viral vector–mediated gene delivery is an attractive procedure for introducing genes into the brain, both for purposes of basic neuroscience research and to develop gene therapy for neurological diseases. Replication-defective adenoviruses possess many features which make them ideal vectors for this purpose—efficiently transducing terminally differentiated cells such as neurons and glial cells, resulting in high levels of transgene expression in vivo. Also, in the absence of anti-adenovirus immunity, these vectors can sustain very long-term transgene expression within the brain parenchyma. This unit provides protocols for the stereotactic injection of adenoviral vectors into the brain, followed by protocols to detect transgene expression or infiltrates of immune cells by immunocytochemistry or immunofluorescence. ELISPOT and neutralizing antibody assay methodologies are provided to quantitate the levels of cellular and humoral immune responses against adenoviruses. Quantitation of adenoviral vector genomes within the rat brain using qPCR is also described. Curr. Protoc. Neurosci. 50:4.24.1–4.24.49. © 2010 by John Wiley & Sons, Inc. PMID:20066657

  14. Potential transfer of extended spectrum β-lactamase encoding gene, blashv18 gene, between Klebsiella pneumoniae in raw foods.

    PubMed

    Jung, Yangjin; Matthews, Karl R

    2016-12-01

    This study investigated the transfer frequency of the extended-spectrum β-lactamase-encoding gene (blaSHV18) among Klebsiella pneumoniae in tryptic soy broth (TSB), pasteurized milk, unpasteurized milk, alfalfa sprouts and chopped lettuce at defined temperatures. All transconjugants were characterized phenotypically and genotypically. KP04(ΔKM) and KP08(ΔKM) isolated from seed sprouts and KP342 were used as recipients in mating experiments with K. pneumoniae ATCC 700603 serving as the donor. In mating experiments, no transconjugants were detected at 4 °C in liquid media or chopped lettuce, but detected in all media tested at 15 °C, 24 °C, and 37 °C. At 24 °C, the transfer of blaSHV18 gene occurred more frequently in alfalfa sprouts (5.15E-04 transconjugants per recipient) and chopped lettuce (3.85E-05) than liquid media (1.08E-05). On chopped lettuce, transconjugants were not detected at day 1 post-mating at 15 °C, but observed on day 2 (1.43E-05). Transconjugants carried the blaSHV18 gene transferred from the donor and the virulence gene harbored by recipient. More importantly, a class 1 integrase gene and resistance to tetracycline, trimethoprim/sulfamethoxazole were co-transferred during mating. These quantitative results suggest that fresh produce exposed to temperature abuse may serve as a competent vehicle for the spread of gene encoding for antibiotic resistance, having a potential negative impact on human health. PMID:27554144

  15. Herpes simplex virus-mediated human hypoxanthine-guanine phosphoribosyltransferase gene transfer into neuronal cells

    SciTech Connect

    Palella, T.D.; Silverman, L.J.; Schroll, C.T.; Homa, F.L.; Levine, M.; Kelley, W.N.

    1988-01-01

    The virtually complete deficiency of the purine salvage enzyme hypoxanthine-guanine phosphoribosyltransferase (HPRT) results in a devastating neurological disease, Lesch-Nyhan syndrome. Transfer of the HPRT gene into fibroblasts and lymphoblasts in vitro and into hematopoietic cells in vivo has been accomplished by other groups with retroviral-derived vectors. It appears to be necessary, however, to transfer the HPRT gene into neuronal cells to correct the neurological dysfunction of this disorder. The neurotropic virus herpes simplex virus type 1 has features that make it suitable for use as a vector to transfer the HPRT gene into neuronal tissue. This report describes the isolation of an HPRT-deficient rat neuroma cell line, designated B103-4C, and the construction of a recombinant herpes simplex virus type 1 that contained human HPRT cDNA. These recombinant viruses were used to infect B103-4C cells. Infected cells expressed HPRT activity which was human in origin.

  16. Adenoviral-Mediated Imaging of Gene Transfer Using a Somatostatin Receptor-Cytosine Deaminase Fusion Protein

    PubMed Central

    Lears, Kimberly A.; Parry, Jesse J.; Andrews, Rebecca; Nguyen, Kim; Wadas, Thaddeus J.; Rogers, Buck E.

    2015-01-01

    Suicide gene therapy is a process by which cells are administered a gene that encodes a protein capable of converting a nontoxic prodrug into an active toxin. Cytosine deaminase (CD) has been widely investigated as a means of suicide gene therapy due to the enzyme’s ability to convert the prodrug 5-fluorocytosine (5-FC) into the toxic compound 5-fluorouracil (5-FU). However, the extent of gene transfer is a limiting factor in predicting therapeutic outcome. The ability to monitor gene transfer, non-invasively, would strengthen the efficiency of therapy. In this regard, we have constructed and evaluated a replication-deficient adenovirus (Ad) containing the human somatostatin receptor subtype 2 (SSTR2) fused with a C-terminal yeast CD gene for the non-invasive monitoring of gene transfer and therapy. The resulting Ad (AdSSTR2-yCD) was evaluated in vitro in breast cancer cells to determine the function of the fusion protein. These studies demonstrated that the both the SSTR2 and yCD were functional in binding assays, conversion assays, and cytotoxicity assays. In vivo studies similarly demonstrated the functionality using conversion assays, biodistribution studies, and small animal positron-emission tomography (PET) imaging studies. In conclusion, the fusion protein has been validated as useful for the non-invasive imaging of yCD expression and will be evaluated in the future for monitoring yCD-based therapy. PMID:25837665

  17. Stable and Efficient Gene Transfer into the Retina Using an HIV-Based Lentiviral Vector

    NASA Astrophysics Data System (ADS)

    Miyoshi, Hiroyuki; Takahashi, Masayo; Gage, Fred H.; Verma, Inder M.

    1997-09-01

    The development of methods for efficient gene transfer to terminally differentiated retinal cells is important to study the function of the retina as well as for gene therapy of retinal diseases. We have developed a lentiviral vector system based on the HIV that can transduce terminally differentiated neurons of the brain in vivo. In this study, we have evaluated the ability of HIV vectors to transfer genes into retinal cells. An HIV vector containing a gene encoding the green fluorescent protein (GFP) was injected into the subretinal space of rat eyes. The GFP gene under the control of the cytomegalovirus promoter was efficiently expressed in both photoreceptor cells and retinal pigment epithelium. However, the use of the rhodopsin promoter resulted in expression predominantly in photoreceptor cells. Most successfully transduced eyes showed that photoreceptor cells in >80% of the area of whole retina expressed the GFP. The GFP expression persisted for at least 12 weeks with no apparent decrease. The efficient gene transfer into photoreceptor cells by HIV vectors will be useful for gene therapy of retinal diseases such as retinitis pigmentosa.

  18. Photoregulation of a phytochrome gene promoter from oat transferred into rice by particle bombardment.

    PubMed Central

    Bruce, W B; Christensen, A H; Klein, T; Fromm, M; Quail, P H

    1989-01-01

    The regulatory photoreceptor phytochrome controls the transcription of its own phy genes in a negative feedback fashion. We have exploited microprojectile-mediated gene transfer to develop a rapid transient expression assay system for the study of DNA sequences involved in the phytochrome-regulated expression of these genes. The 5'-flanking sequence and part of the structural region of an oat phy gene have been fused to a reporter coding sequence (chloramphenicol acetyltransferase, CAT) and introduced into intact darkgrown seedlings by using high-velocity microprojectiles. Expression is assayable in less than 24 hr from bombardment. The introduced oat phy-CAT fusion gene is expressed and down-regulated by white light in barley, rice, and oat, whereas no expression is detected in three dicots tested, tobacco, cucumber, and Arabidopsis thaliana. In bombarded rice shoots, red/far-red light-reversible repression of expression of the heterologous oat phy-CAT gene shows that it is regulated by phytochrome in a manner parallel to that of the endogenous rice phy genes. These data indicate that the transduction pathway components and promoter sequences involved in autoregulation of phy expression have been evolutionarily conserved between oat and rice. The experiments show the feasibility of using high-velocity microprojectile-mediated gene transfer for the rapid analysis of light-controlled monocot gene promoters in monocot tissues that until now have been recalcitrant to such studies. Images PMID:2602370

  19. Horizontal Transfer of Plasmid-Mediated Cephalosporin Resistance Genes in the Intestine of Houseflies (Musca domestica).

    PubMed

    Fukuda, Akira; Usui, Masaru; Okubo, Torahiko; Tamura, Yutaka

    2016-06-01

    Houseflies are a mechanical vector for various types of bacteria, including antimicrobial-resistant bacteria (ARB). If the intestine of houseflies is a suitable site for the transfer of antimicrobial resistance genes (ARGs), houseflies could also serve as a biological vector for ARB. To clarify whether cephalosporin resistance genes are transferred efficiently in the housefly intestine, we compared with conjugation experiments in vivo (in the intestine) and in vitro by using Escherichia coli with eight combinations of four donor and two recipient strains harboring plasmid-mediated cephalosporin resistance genes and chromosomal-encoded rifampicin resistance genes, respectively. In the in vivo conjugation experiment, houseflies ingested donor strains for 6 hr and then recipient strains for 3 hr, and 24 hr later, the houseflies were surface sterilized and analyzed. In vitro conjugation experiments were conducted using the broth-mating method. In 3/8 combinations, the in vitro transfer frequency (Transconjugants/Donor) was ≥1.3 × 10(-4); the in vivo transfer rates of cephalosporin resistance genes ranged from 2.0 × 10(-4) to 5.7 × 10(-5). Moreover, cephalosporin resistance genes were transferred to other species of enteric bacteria of houseflies such as Achromobacter sp. and Pseudomonas fluorescens. These results suggest that houseflies are not only a mechanical vector for ARB but also a biological vector for the occurrence of new ARB through the horizontal transfer of ARGs in their intestine. PMID:26683492

  20. Influence of Host Interleukin-10 Polymorphisms on Development of Traveler's Diarrhea Due to Heat-Labile Enterotoxin-Producing Escherichia coli in Travelers from the United States Who Are Visiting Mexico▿

    PubMed Central

    Flores, Jose; DuPont, Herbert L.; Lee, Stephanie A.; Belkind-Gerson, Jaime; Paredes, Mercedes; Mohamed, Jamal A.; Armitige, Lisa Y.; Guo, Dong-Chuan; Okhuysen, Pablo C.

    2008-01-01

    Up to 60% of U.S. visitors to Mexico develop traveler's diarrhea (TD), mostly due to enterotoxigenic Escherichia coli (ETEC) strains that produce heat-labile (LT) and/or heat-stable (ST) enterotoxins. Distinct single-nucleotide polymorphisms (SNPs) within the interleukin-10 (IL-10) promoter have been associated with high, intermediate, or low production of IL-10. We conducted a prospective study to investigate the association of SNPs in the IL-10 promoter and the occurrence of TD in ETEC LT-exposed travelers. Sera from U.S. travelers to Mexico collected on arrival and departure were studied for ETEC LT seroconversion by using cholera toxin as the antigen. Pyrosequencing was performed to genotype IL-10 SNPs. Stools from subjects who developed diarrhea were also studied for other enteropathogens. One hundred twenty-one of 569 (21.3%) travelers seroconverted to ETEC LT, and among them 75 (62%) developed diarrhea. Symptomatic seroconversion was more commonly seen in subjects who carried a genotype producing high levels of IL-10; it was seen in 83% of subjects with the GG genotype versus 54% of subjects with the AA genotype at IL-10 gene position −1082 (P, 0.02), in 71% of those with the CC genotype versus 33% of those with the TT genotype at position −819 (P, 0.005), and in 71% of those with the CC genotype versus 38% of those with the AA genotype at position −592 (P, 0.02). Travelers with the GCC haplotype were more likely to have symptomatic seroconversion than those with the ATA haplotype (71% versus 38%; P, 0.002). Travelers genetically predisposed to produce high levels of IL-10 were more likely to experience symptomatic ETEC TD. PMID:18579697

  1. Lack of Interleukin-10-Mediated Anti-Inflammatory Signals and Upregulated Interferon Gamma Production Are Linked to Increased Intestinal Epithelial Cell Apoptosis in Pathogenic Simian Immunodeficiency Virus Infection

    PubMed Central

    Pan, Diganta; Kenway-Lynch, Carys S.; Lala, Wendy; Veazey, Ronald S.; Lackner, Andrew A.; Das, Arpita

    2014-01-01

    ABSTRACT Interleukin-10 (IL-10) is an immunomodulatory cytokine that is important for maintenance of epithelial cell (EC) survival and anti-inflammatory responses (AIR). The majority of HIV infections occur through the mucosal route despite mucosal epithelium acting as a barrier to human immunodeficiency virus (HIV). Therefore, understanding the role of IL-10 in maintenance of intestinal homeostasis during HIV infection is of interest for better characterization of the pathogenesis of HIV-mediated enteropathy. We demonstrated here changes in mucosal IL-10 signaling during simian immunodeficiency virus (SIV) infection in rhesus macaques. Disruption of the epithelial barrier was manifested by EC apoptosis and loss of the tight-junction protein ZO-1. Multiple cell types, including a limited number of ECs, produced IL-10. SIV infection resulted in increased levels of IL-10; however, this was associated with increased production of mucosal gamma interferon (IFN-γ) and tumor necrosis factor alpha (TNF-α), suggesting that IL-10 was not able to regulate AIR. This observation was supported by the downregulation of STAT3, which is necessary to inhibit production of IFN-γ and TNF-α, and the upregulation of SOCS1 and SOCS3, which are important regulatory molecules in the IL-10-mediated AIR. We also observed internalization of the IL-10 receptor (IL-10R) in mucosal lymphocytes, which could limit cellular availability of IL-10 for signaling and contribute to the loss of a functional AIR. Collectively, these findings demonstrate that internalization of IL-10R with the resultant impact on IL-10 signaling and dysregulation of the IL-10-mediated AIR might play a crucial role in EC damage and subsequent SIV/HIV pathogenesis. IMPORTANCE Interleukin-10 (IL-10), an important immunomodulatory cytokine plays a key role to control inflammatory function and homeostasis of the gastrointestinal mucosal immune system. Despite recent advancements in the study of IL-10 and its role in HIV

  2. Improved retroviral suicide gene transfer in colon cancer cell lines after cell synchronization with methotrexate

    PubMed Central

    2011-01-01

    Background Cancer gene therapy by retroviral vectors is mainly limited by the level of transduction. Retroviral gene transfer requires target cell division. Cell synchronization, obtained by drugs inducing a reversible inhibition of DNA synthesis, could therefore be proposed to precondition target cells to retroviral gene transfer. We tested whether drug-mediated cell synchronization could enhance the transfer efficiency of a retroviral-mediated gene encoding herpes simplex virus thymidine kinase (HSV-tk) in two colon cancer cell lines, DHDK12 and HT29. Methods Synchronization was induced by methotrexate (MTX), aracytin (ara-C) or aphidicolin. Gene transfer efficiency was assessed by the level of HSV-TK expression. Transduced cells were driven by ganciclovir (GCV) towards apoptosis that was assessed using annexin V labeling by quantitative flow cytometry. Results DHDK12 and HT29 cells were synchronized in S phase with MTX but not ara-C or aphidicolin. In synchronized DHDK12 and HT29 cells, the HSV-TK transduction rates were 2 and 1.5-fold higher than those obtained in control cells, respectively. Furthermore, the rate of apoptosis was increased two-fold in MTX-treated DHDK12 cells after treatment with GCV. Conclusions Our findings indicate that MTX-mediated synchronization of target cells allowed a significant improvement of retroviral HSV-tk gene transfer, resulting in an increased cell apoptosis in response to GCV. Pharmacological control of cell cycle may thus be a useful strategy to optimize the efficiency of retroviral-mediated cancer gene therapy. PMID:21970612

  3. Transferring Gus gene into intact rice cells by low energy ion beam

    NASA Astrophysics Data System (ADS)

    Zengliang, Yu; Jianbo, Yang; Yuejin, Wu; Beijiu, Cheng; Jianjun, He; Yuping, Huo

    1993-06-01

    A new technique of transferring genes by low energy ion beam has been reported in this paper. The Gus and CAT (chloramphenicol acetyltransferase) genes, as "foreign" genetic materials, were introduced into the suspension cells and ripe embryos or rice by implantation of 20-30 keV Ar + at doses ranging from 1 × 10 15 to 4 × 10 15 ions/cm 2. The activities of CAT and Gus were detected in the cells and embryos after several weeks. The results indicate that the transfer was a success.

  4. Efficient Gene Transfer and Targeted Mutagenesis in Fusobacterium nucleatum

    PubMed Central

    Haake, Susan Kinder; Yoder, Sean; Gerardo, Sharon Hunt

    2006-01-01

    Fusobacterium nucleatum is a Gram-negative anaerobe important in dental biofilm ecology and infectious diseases with significant societal impact. The lack of efficient genetic systems has hampered molecular analyses in this microorganism. We previously reported construction of a shuttle plasmid, pHS17, using the native fusobacterial plasmid pFN1 and an erythromycin resistance cassette. However, the host range of pHS17 was restricted to F. nucleatum, ATCC 10953 and the transformation efficiency was limited. This study was undertaken to improve genetic systems for molecular analysis in F. nucleatum. We identified a second F. nucleatum strain, ATCC 23726, which is transformed with improved efficiency compared to ATCC 10953. Two novel second generation pFN1-based shuttle plasmids, pHS23 and pHS30, were developed and enable transformation of ATCC 23726 at 6.2 x 104 and 1.5 x 106 transformants/microgram of plasmid DNA, respectively. The transformation efficiency of pHS30, which harbors a catP gene conferring resistance to chloramphenicol, was more than 1,000-fold greater than that of pHS17. The improved transformation efficiency facilitated disruption of the chromosomal rnr gene using a suicide plasmid pHS19, the first demonstration of targeted mutagenesis in F. nucleatum. These results provide significant advances in the development of systems for molecular analysis in F. nucleatum. PMID:16115683

  5. Nano-vectors for efficient liver specific gene transfer

    PubMed Central

    Pathak, Atul; Vyas, Suresh P; Gupta, Kailash C

    2008-01-01

    Recent progress in nanotechnology has triggered the site specific drug/gene delivery research and gained wide acknowledgment in contemporary DNA therapeutics. Amongst various organs, liver plays a crucial role in various body functions and in addition, the site is a primary location of metastatic tumor growth. In past few years, a plethora of nano-vectors have been developed and investigated to target liver associated cells through receptor mediated endocytosis. This emerging paradigm in cellular drug/gene delivery provides promising approach to eradicate genetic as well as acquired diseases affecting the liver. The present review provides a comprehensive overview of potential of various delivery systems, viz., lipoplexes, liposomes, polyplexes, nanoparticles and so forth to selectively relocate foreign therapeutic DNA into liver specific cell type via the receptor mediated endocytosis. Various receptors like asialoglycoprotein receptors (ASGP-R) provide unique opportunity to target liver parenchymal cells. The results obtained so far reveal tremendous promise and offer enormous options to develop novel DNA-based pharmaceuticals for liver disorders in near future. PMID:18488414

  6. In situ gene transfer and suicide gene therapy of gastric cancer induced by N-ethyl-N'-nitro-N-nitrosoguanidine in dogs.

    PubMed

    Matsukura, N; Hoshino, A; Igarashi, T; Hasegawa, H; Okino, T; Onda, M; Iijima, O; Akiyama, K; Goto, T; Takubo, K; Suzuki, S; Shimada, T

    1999-09-01

    Gene therapy could potentially revolutionize the treatment of gastrointestinal (GI) tract cancer. The aim of this study was to establish a practical method of gene transfer which would be applicable to human gastric cancer. Retrovirus or/and adenovirus vectors carrying the lacZ marker gene were transferred in situ by needle through an endoscopic biopsy channel into primary gastric cancer in six male beagle dogs that had been treated with N-ethyl-N'-nitro-N-nitrosoguanidine (ENNG). In addition, an adenovirus vector carrying the herpes simplex virus thymidine kinase (Ad.CAGHSV-TK) gene was introduced in situ into cancer tissues in the stomach of three dogs, and the animals were treated with intravenous ganciclovir (GCV). Retrovirus-producing cells which expressed the lacZ gene were specifically localized to the injection site in the stomach. The lacZ gene was more widely transferred into the tumor by the adenovirus vector than by retrovirus-producing cells. Improvement of the needle used for gene transfer and the use of multiple injections per tumor led to more diffuse transfer of the vector into the tumor. The Ad.CAGlacZ gene was also transferred into regional lymph nodes of the stomach. Moderate to diffuse degeneration of the primary cancer tissues of the stomach was found after Ad.CAGHSV-TK/GCV gene therapy. Moreover, almost complete tissue degeneration was observed in the regional lymph nodes of the stomach. An adverse effect of HSV-TK/GCV gene therapy was acute hepatotoxicity, which was not found after Ad.CAGlacZ gene transfer, but was found after high-titer Ad.CAGHSV-TK gene transfer followed by GCV. These findings suggest that in situ gene transfer of a suicide gene followed by prodrug treatment may be applicable not only to primary tumors, but also to lymph node metastases of gastric cancer, though further study of both beneficial and adverse effects is required before clinical usage. PMID:10551335

  7. Regulatory and ethical issues for phase I in utero gene transfer studies.

    PubMed

    Strong, Carson

    2011-11-01

    Clinical gene transfer research has involved adult and child subjects, and it is expected that gene transfer in fetal subjects will occur in the future. Some genetic diseases have serious adverse effects on the fetus before birth, and there is hope that prenatal gene therapy could prevent such disease progression. Research in animal models of prenatal gene transfer is actively being pursued. The prospect of human phase I in utero gene transfer studies raises important regulatory and ethical issues. One issue not previously addressed arises in applying U.S. research regulations to such studies. Specifically, current regulations state that research involving greater than minimal risk to the fetus and no prospect of direct benefit to the fetus or pregnant woman is not permitted. Phase I studies will involve interventions such as needle insertions through the uterus, which carry risks to the fetus including spontaneous abortion and preterm birth. It is possible that these risks will be regarded as exceeding minimal. Also, some regard the probability of therapeutic benefit in phase I studies to be so low that these studies do not satisfy the regulatory requirement that they "hold out the prospect of direct benefit" to subjects. On the basis of these considerations, investigators and institutional review boards might reasonably conclude that some phase I in utero studies are not to be permitted. This paper identifies considerations that are relevant to such judgments and explores ethically acceptable ways in which phase I studies can be designed so that they are permitted by the regulations. PMID:21846200

  8. Peptide nanofibrils boost retroviral gene transfer and provide a rapid means for concentrating viruses

    NASA Astrophysics Data System (ADS)

    Yolamanova, Maral; Meier, Christoph; Shaytan, Alexey K.; Vas, Virag; Bertoncini, Carlos W.; Arnold, Franziska; Zirafi, Onofrio; Usmani, Shariq M.; Müller, Janis A.; Sauter, Daniel; Goffinet, Christine; Palesch, David; Walther, Paul; Roan, Nadia R.; Geiger, Hartmut; Lunov, Oleg; Simmet, Thomas; Bohne, Jens; Schrezenmeier, Hubert; Schwarz, Klaus; Ständker, Ludger; Forssmann, Wolf-Georg; Salvatella, Xavier; Khalatur, Pavel G.; Khokhlov, Alexei R.; Knowles, Tuomas P. J.; Weil, Tanja; Kirchhoff, Frank; Münch, Jan

    2013-02-01

    Inefficient gene transfer and low virion concentrations are common limitations of retroviral transduction. We and others have previously shown that peptides derived from human semen form amyloid fibrils that boost retroviral gene delivery by promoting virion attachment to the target cells. However, application of these natural fibril-forming peptides is limited by moderate efficiencies, the high costs of peptide synthesis, and variability in fibril size and formation kinetics. Here, we report the development of nanofibrils that self-assemble in aqueous solution from a 12-residue peptide, termed enhancing factor C (EF-C). These artificial nanofibrils enhance retroviral gene transfer substantially more efficiently than semen-derived fibrils or other transduction enhancers. Moreover, EF-C nanofibrils allow the concentration of retroviral vectors by conventional low-speed centrifugation, and are safe and effective, as assessed in an ex vivo gene transfer study. Our results show that EF-C fibrils comprise a highly versatile, convenient and broadly applicable nanomaterial that holds the potential to significantly facilitate retroviral gene transfer in basic research and clinical applications.

  9. Different gene transfer methods at the very early, early, late and whole embryonic stages in chicken.

    PubMed

    Gong, Ping; Yang, Y P; Yang, Y; Feng, Yan P; Li, S J; Peng, Xiu L; Gong, Y Z

    2012-12-01

    New technologies in gene transfer combined with experimental embryology make the chicken embryo an excellent model system for gene function studies. The techniques of in ovo electroporation, in vitro culture for ex ovo electroporation and retrovirus-mediated gene transfer have already been fully developed in chicken. Yet to our knowledge, there are no definite descriptions on the features and application scopes of these techniques. The survival rates of different in vitro culture methods were compared and the EGFP expression areas of different gene transfer techniques were explored. It was that the optimal timings of removing embryo for EC culture and Petri dish system was at E1.5 and E2.5, respectively; and optimal timing of injecting retrovirus is at E0. Results indicated that the EC culture, in ovo electroporation, the Petri dish system and retrovirus-mediated method are, respectively, suitable for the very early, early, late and whole embryonic stages in chicken. Comparison of different gene transfer methods and establishment of optimal timings are expected to provide a better choice of the efficient method for a particular experiment. PMID:23134602

  10. Bacteriophage Mediates Efficient Gene Transfer in Combination with Conventional Transfection Reagents

    PubMed Central

    Donnelly, Amanda; Yata, Teerapong; Bentayebi, Kaoutar; Suwan, Keittisak; Hajitou, Amin

    2015-01-01

    The development of commercially available transfection reagents for gene transfer applications has revolutionized the field of molecular biology and scientific research. However, the challenge remains in ensuring that they are efficient, safe, reproducible and cost effective. Bacteriophage (phage)-based viral vectors have the potential to be utilized for general gene transfer applications within research and industry. Yet, they require adaptations in order to enable them to efficiently enter cells and overcome mammalian cellular barriers, as they infect bacteria only; furthermore, limited progress has been made at increasing their efficiency. The production of a novel hybrid nanocomplex system consisting of two different nanomaterial systems, phage vectors and conventional transfection reagents, could overcome these limitations. Here we demonstrate that the combination of cationic lipids, cationic polymers or calcium phosphate with M13 bacteriophage-derived vectors, engineered to carry a mammalian transgene cassette, resulted in increased cellular attachment, entry and improved transgene expression in human cells. Moreover, addition of a targeting ligand into the nanocomplex system, through genetic engineering of the phage capsid further increased gene expression and was effective in a stable cell line generation application. Overall, this new hybrid nanocomplex system (i) provides enhanced phage-mediated gene transfer; (ii) is applicable for laboratory transfection processes and (iii) shows promise within industry for large-scale gene transfer applications. PMID:26670247

  11. Collective evolution of cyanobacteria and cyanophages mediated by horizontal gene transfer

    NASA Astrophysics Data System (ADS)

    Shih, Hong-Yan; Rogers, Tim; Goldenfeld, Nigel

    We describe a model for how antagonistic predator-prey coevolution can lead to mutualistic adaptation to an environment, as a result of horizontal gene transfer. Our model is a simple description of ecosystems such as marine cyanobacteria and their predator cyanophages, which carry photosynthesis genes. These genes evolve more rapidly in the virosphere than the bacterial pan-genome, and thus the bacterial population could potentially benefit from phage predation. By modeling both the barrier to predation and horizontal gene transfer, we study this balance between individual sacrifice and collective benefits. The outcome is an emergent mutualistic coevolution of improved photosynthesis capability, benefiting both bacteria and phage. This form of multi-level selection can contribute to niche stratification in the cyanobacteria-phage ecosystem. This work is supported in part by a cooperative agreement with NASA, Grant NNA13AA91A/A0018.

  12. Targeting the urokinase plasminogen activator receptor enhances gene transfer to human airway epithelia

    PubMed Central

    Drapkin, Paola T.; O’Riordan, Catherine R.; Yi, Su Min; Chiorini, John A.; Cardella, Jonathan; Zabner, Joseph; Welsh, Michael J.

    2000-01-01

    Developing gene therapy for cystic fibrosis has been hindered by limited binding and endocytosis of vectors by human airway epithelia. Here we show that the apical membrane of airway epithelia express the urokinase plasminogen activator receptor (uPAR). Urokinase plasminogen activator (uPA), or a 7-residue peptide derived from this protein (u7-peptide), bound the receptor and stimulated apical endocytosis. Both ligands enhanced gene transfer by nonspecifically bound adenovirus and adeno-associated virus vectors and by a modified adenovirus vector that had been coupled to the u7-peptide. These data provide the first evidence that targeting an apical receptor can circumvent the two most important barriers to gene transfer in airway epithelia. Thus, the uPA/uPAR system may offer significant advantages for delivering genes and other pharmaceuticals to airway epithelia. PMID:10712430

  13. Targeting the urokinase plasminogen activator receptor enhances gene transfer to human airway epithelia.

    PubMed

    Drapkin, P T; O'Riordan, C R; Yi, S M; Chiorini, J A; Cardella, J; Zabner, J; Welsh, M J

    2000-03-01

    Developing gene therapy for cystic fibrosis has been hindered by limited binding and endocytosis of vectors by human airway epithelia. Here we show that the apical membrane of airway epithelia express the urokinase plasminogen activator receptor (uPAR). Urokinase plasminogen activator (uPA), or a 7-residue peptide derived from this protein (u7-peptide), bound the receptor and stimulated apical endocytosis. Both ligands enhanced gene transfer by nonspecifically bound adenovirus and adeno-associated virus vectors and by a modified adenovirus vector that had been coupled to the u7-peptide. These data provide the first evidence that targeting an apical receptor can circumvent the two most important barriers to gene transfer in airway epithelia. Thus, the uPA/uPAR system may offer significant advantages for delivering genes and other pharmaceuticals to airway epithelia. PMID:10712430

  14. Rate of gene transfer from mitochondria to nucleus: effects of cytoplasmic inheritance system and intensity of intracellular competition.

    PubMed

    Yamauchi, Atsushi

    2005-11-01

    Endosymbiotic theory states that mitochondria originated as bacterial intracellular symbionts, the size of the mitochondrial genome gradually reducing over a long period owing to, among other things, gene transfer from the mitochondria to the nucleus. Such gene transfer was observed in more genes in animals than in plants, implying a higher transfer rate of animals. The evolution of gene transfer may have been affected by an intensity of intracellular competition among organelle strains and the organelle inheritance system of the organism concerned. This article reveals a relationship between those factors and the gene transfer rate from organelle to nuclear genomes, using a mathematical model. Mutant mitochondria that lose a certain gene by deletion are considered to replicate more rapidly than normal ones, resulting in an advantage in intracellular competition. If the competition is intense, heteroplasmic individuals possessing both types of mitochondria change to homoplasmic individuals including mutant mitochondria only, with high probability. According to the mathematical model, it was revealed that the rate of gene transfer from mitochondria to the nucleus can be affected by three factors, the intensity of intracellular competition, the probability of paternal organelle transmission, and the effective population size. The gene transfer rate tends to increase with decreasing intracellular competition, increasing paternal organelle transmission, and decreasing effective population size. Intense intracellular competition tends to suppress gene transfer because it is likely to exclude mutant mitochondria that lose the essential gene due to the production of lethal individuals. PMID:16079242

  15. Operon Formation is Driven by Co-Regulation and Not by Horizontal Gene Transfer

    SciTech Connect

    Price, Morgan N.; Huang, Katherine H.; Arkin, Adam P.; Alm, Eric J.

    2005-04-12

    Although operons are often subject to horizontal gene transfer (HGT), non-HGT genes are particularly likely to be in operons. To resolve this apparent discrepancy and to determine whether HGT is involved in operon formation, we examined the evolutionary history of the genes and operons in Escherichia coli K12. We show that genes that have homologs in distantly related bacteria but not in close relatives of E. coli (indicating HGTi) form new operons at about the same rates as native genes. Furthermore, genes in new operons are no more likely than other genes to have phylogenetic trees that are inconsistent with the species tree. In contrast, essential genes and ubiquitous genes without paralogs (genes believed to undergo HGT rarely) often form new operons. We conclude that HGT is not associated with operon formation, but instead promotes the prevalence of pre-existing operons. To explain operon formation, we propose that new operons reduce the amount of regulatory information required to specify optimal expression patterns. Consistent with this hypothesis, operons have greater amounts of conserved regulatory sequences than do individually transcribed genes.

  16. Inferring gene duplications, transfers and losses can be done in a discrete framework.

    PubMed

    Ranwez, Vincent; Scornavacca, Celine; Doyon, Jean-Philippe; Berry, Vincent

    2016-06-01

    In the field of phylogenetics, the evolutionary history of a set of organisms is commonly depicted by a species tree-whose internal nodes represent speciation events-while the evolutionary history of a gene family is depicted by a gene tree-whose internal nodes can also represent macro-evolutionary events such as gene duplications and transfers. As speciation events are only part of the events shaping a gene history, the topology of a gene tree can show incongruences with that of the corresponding species tree. These incongruences can be used to infer the macro-evolutionary events undergone by the gene family. This is done by embedding the gene tree inside the species tree and hence providing a reconciliation of those trees. In the past decade, several parsimony-based methods have been developed to infer such reconciliations, accounting for gene duplications ([Formula: see text]), transfers ([Formula: see text]) and losses ([Formula: see text]). The main contribution of this paper is to formally prove an important assumption implicitly made by previous works on these reconciliations, namely that solving the (maximum) parsimony [Formula: see text] reconciliation problem in the discrete framework is equivalent to finding a most parsimonious [Formula: see text] scenario in the continuous framework. In the process, we also prove several intermediate results that are useful on their own and constitute a theoretical toolbox that will likely facilitate future theoretical contributions in the field. PMID:26337177

  17. Horizontal Transfer of a Nitrate Assimilation Gene Cluster and Ecological Transitions in Fungi: A Phylogenetic Study

    PubMed Central

    Slot, Jason C.; Hibbett, David S.

    2007-01-01

    High affinity nitrate assimilation genes in fungi occur in a cluster (fHANT-AC) that can be coordinately regulated. The clustered genes include nrt2, which codes for a high affinity nitrate transporter; euknr, which codes for nitrate reductase; and NAD(P)H-nir, which codes for nitrite reductase. Homologs of genes in the fHANT-AC occur in other eukaryotes and prokaryotes, but they have only been found clustered in the oomycete Phytophthora (heterokonts). We performed independent and concatenated phylogenetic analyses of homologs of all three genes in the fHANT-AC. Phylogenetic analyses limited to fungal sequences suggest that the fHANT-AC has been transferred horizontally from a basidiomycete (mushrooms and smuts) to an ancestor of the ascomycetous mold Trichoderma reesei. Phylogenetic analyses of sequences from diverse eukaryotes and eubacteria, and cluster structure, are consistent with a hypothesis that the fHANT-AC was assembled in a lineage leading to the oomycetes and was subsequently transferred to the Dikarya (Ascomycota+Basidiomycota), which is a derived fungal clade that includes the vast majority of terrestrial fungi. We propose that the acquisition of high affinity nitrate assimilation contributed to the success of Dikarya on land by allowing exploitation of nitrate in aerobic soils, and the subsequent transfer of a complete assimilation cluster improved the fitness of T. reesei in a new niche. Horizontal transmission of this cluster of functionally integrated genes supports the “selfish operon” hypothesis for maintenance of gene clusters. PMID:17971860

  18. Phylogenomic analysis demonstrates a pattern of rare and ancient horizontal gene transfer between plants and fungi.

    PubMed

    Richards, Thomas A; Soanes, Darren M; Foster, Peter G; Leonard, Guy; Thornton, Christopher R; Talbot, Nicholas J

    2009-07-01

    Horizontal gene transfer (HGT) describes the transmission of genetic material across species boundaries and is an important evolutionary phenomenon in the ancestry of many microbes. The role of HGT in plant evolutionary history is, however, largely unexplored. Here, we compare the genomes of six plant species with those of 159 prokaryotic and eukaryotic species and identify 1689 genes that show the highest similarity to corresponding genes from fungi. We constructed a phylogeny for all 1689 genes identified and all homolog groups available from the rice (Oryza sativa) genome (3177 gene families) and used these to define 14 candidate plant-fungi HGT events. Comprehensive phylogenetic analyses of these 14 data sets, using methods that account for site rate heterogeneity, demonstrated support for nine HGT events, demonstrating an infrequent pattern of HGT between plants and fungi. Five HGTs were fungi-to-plant transfers and four were plant-to-fungi HGTs. None of the fungal-to-plant HGTs involved angiosperm recipients. These results alter the current view of organismal barriers to HGT, suggesting that phagotrophy, the consumption of a whole cell by another, is not necessarily a prerequisite for HGT between eukaryotes. Putative functional annotation of the HGT candidate genes suggests that two fungi-to-plant transfers have added phenotypes important for life in a soil environment. Our study suggests that genetic exchange between plants and fungi is exceedingly rare, particularly among the angiosperms, but has occurred during their evolutionary history and added important metabolic traits to plant lineages. PMID:19584142

  19. Preventing High Fat Diet-induced Obesity and Improving Insulin Sensitivity through Neuregulin 4 Gene Transfer

    PubMed Central

    Ma, Yongjie; Gao, Mingming; Liu, Dexi

    2016-01-01

    Neuregulin 4 (NRG4), an epidermal growth factor-like signaling molecule, plays an important role in cell-to-cell communication during tissue development. Its function to regulate energy metabolism has recently been reported. This current study was designed to assess the preventive and therapeutic effects of NRG4 overexpression on high fat diet (HFD)-induced obesity. Using the hydrodynamic gene transfer method, we demonstrate that Nrg4 gene transfer in mice suppressed the development of diet-induced obesity, but did not affect pre-existing adiposity and body weight in obese mice. Nrg4 gene transfer curbed HFD-induced hepatic steatosis by inhibiting lipogenesis and PPARγ-mediated lipid storage. Concurrently, overexpression of NRG4 reduced chronic inflammation in both preventive and treatment studies, evidenced by lower mRNA levels of macrophage marker genes including F4/80, Cd68, Cd11b, Cd11c, and macrophage chemokine Mcp1, resulting in improved insulin sensitivity. Collectively, these results demonstrate that overexpression of the Nrg4 gene by hydrodynamic gene delivery prevents HFD-induced weight gain and fatty liver, alleviates obesity-induced chronic inflammation and insulin resistance, and supports the health benefits of NRG4 in managing obesity and obesity-associated metabolic disorders. PMID:27184920

  20. Preventing High Fat Diet-induced Obesity and Improving Insulin Sensitivity through Neuregulin 4 Gene Transfer.

    PubMed

    Ma, Yongjie; Gao, Mingming; Liu, Dexi

    2016-01-01

    Neuregulin 4 (NRG4), an epidermal growth factor-like signaling molecule, plays an important role in cell-to-cell communication during tissue development. Its function to regulate energy metabolism has recently been reported. This current study was designed to assess the preventive and therapeutic effects of NRG4 overexpression on high fat diet (HFD)-induced obesity. Using the hydrodynamic gene transfer method, we demonstrate that Nrg4 gene transfer in mice suppressed the development of diet-induced obesity, but did not affect pre-existing adiposity and body weight in obese mice. Nrg4 gene transfer curbed HFD-induced hepatic steatosis by inhibiting lipogenesis and PPARγ-mediated lipid storage. Concurrently, overexpression of NRG4 reduced chronic inflammation in both preventive and treatment studies, evidenced by lower mRNA levels of macrophage marker genes including F4/80, Cd68, Cd11b, Cd11c, and macrophage chemokine Mcp1, resulting in improved insulin sensitivity. Collectively, these results demonstrate that overexpression of the Nrg4 gene by hydrodynamic gene delivery prevents HFD-induced weight gain and fatty liver, alleviates obesity-induced chronic inflammation and insulin resistance, and supports the health benefits of NRG4 in managing obesity and obesity-associated metabolic disorders. PMID:27184920

  1. Gene Transfer and the Reconstruction of Life's Early History from Genomic Data

    NASA Astrophysics Data System (ADS)

    Gogarten, J. Peter; Fournier, Gregory; Zhaxybayeva, Olga

    2008-03-01

    The metaphor of the unique and strictly bifurcating tree of life, suggested by Charles Darwin, needs to be replaced (or at least amended) to reflect and include processes that lead to the merging of and communication between independent lines of descent. Gene histories include and reflect processes such as gene transfer, symbioses and lineage fusion. No single molecule can serve as a proxy for the tree of life. Individual gene histories can be reconstructed from the growing molecular databases containing sequence and structural information. With some simplifications these gene histories can be represented by furcating trees; however, merging these gene histories into web-like organismal histories, including the transfer of metabolic pathways and cell biological innovations from now-extinct lineages, has yet to be accomplished. Because of these difficulties in interpreting the record retained in molecular sequences, correlations with biochemical fossils and with the geological record need to be interpreted with caution. Advances to detect and pinpoint transfer events promise to untangle at least a few of the intertwined histories of individual genes within organisms and trace them to the organismal ancestors. Furthermore, analysis of the shape of molecular phylogenetic trees may point towards organismal radiations that might reflect early mass extinction events that occurred on a planetary scale.

  2. Gene Transfer and the Reconstruction of Life's Early History from Genomic Data

    NASA Astrophysics Data System (ADS)

    Gogarten, J. Peter; Fournier, Gregory; Zhaxybayeva, Olga

    The metaphor of the unique and strictly bifurcating tree of life, suggested by Charles Darwin, needs to be replaced (or at least amended) to reflect and include processes that lead to the merging of and communication between independent lines of descent. Gene histories include and reflect processes such as gene transfer, symbioses and lineage fusion. No single molecule can serve as a proxy for the tree of life. Individual gene histories can be reconstructed from the growing molecular databases containing sequence and structural information. With some simplifications these gene histories can be represented by furcating trees; however, merging these gene histories into web-like organismal histories, including the transfer of metabolic pathways and cell biological innovations from now-extinct lineages, has yet to be accomplished. Because of these difficulties in interpreting the record retained in molecular sequences, correlations with biochemical fossils and with the geological record need to be interpreted with caution. Advances to detect and pinpoint transfer events promise to untangle at least a few of the intertwined histories of individual genes within organisms and trace them to the organismal ancestors. Furthermore, analysis of the shape of molecular phylogenetic trees may point towards organismal radiations that might reflect early mass extinction events that occurred on a planetary scale.

  3. AAV Vectors for Cardiac Gene Transfer: Experimental Tools and Clinical Opportunities

    PubMed Central

    Pacak, Christina A; Byrne, Barry J

    2011-01-01

    Since the first demonstration of in vivo gene transfer into myocardium there have been a series of advancements that have driven the evolution of cardiac gene delivery from an experimental tool into a therapy currently at the threshold of becoming a viable clinical option. Innovative methods have been established to address practical challenges related to tissue-type specificity, choice of delivery vehicle, potency of the delivered material, and delivery route. Most importantly for therapeutic purposes, these strategies are being thoroughly tested to ensure safety of the delivery system and the delivered genetic material. This review focuses on the development of recombinant adeno-associated virus (rAAV) as one of the most valuable cardiac gene transfer agents available today. Various forms of rAAV have been used to deliver “pre-event” cardiac protection and to temper the severity of hypertrophy, cardiac ischemia, or infarct size. Adeno-associated virus (AAV) vectors have also been functional delivery tools for cardiac gene expression knockdown studies and successfully improving the cardiac aspects of several metabolic and neuromuscular diseases. Viral capsid manipulations along with the development of tissue-specific and regulated promoters have greatly increased the utility of rAAV-mediated gene transfer. Important clinical studies are currently underway to evaluate AAV-based cardiac gene delivery in humans. PMID:21792180

  4. Horizontal gene transfer and nucleotide compositional anomaly in large DNA viruses

    PubMed Central

    Monier, Adam; Claverie, Jean-Michel; Ogata, Hiroyuki

    2007-01-01

    Background DNA viruses have a wide range of genome sizes (5 kb up to 1.2 Mb, compared to 0.16 Mb to 1.5 Mb for obligate parasitic bacteria) that do not correlate with their virulence or the taxonomic distribution of their hosts. The reasons for such large variation are unclear. According to the traditional view of viruses as gifted "gene pickpockets", large viral genome sizes could originate from numerous gene acquisitions from their hosts. We investigated this hypothesis by studying 67 large DNA viruses with genome sizes larger than 150 kb, including the recently characterized giant mimivirus. Given that horizontally transferred DNA often have anomalous nucleotide compositions differing from the rest of the genome, we conducted a detailed analysis of the inter- and intra-genome compositional properties of these viruses. We then interpreted their compositional heterogeneity in terms of possible causes, including strand asymmetry, gene function/expression, and horizontal transfer. Results We first show that the global nucleotide composition and nucleotide word usage of viral genomes are species-specific and distinct from those of their hosts. Next, we identified compositionally anomalous (cA) genes in viral genomes, using a method based on Bayesian inference. The proportion of cA genes is highly variable across viruses and does not exhibit a significant correlation with genome size. The vast majority of the cA genes were of unknown function, lacking homologs in the databases. For genes with known homologs, we found a substantial enrichment of cA genes in specific functional classes for some of the viruses. No significant association was found between cA genes and compositional strand asymmetry. A possible exogenous origin for a small fraction of the cA genes could be confirmed by phylogenetic reconstruction. Conclusion At odds with the traditional dogma, our results argue against frequent genetic transfers to large DNA viruses from their modern hosts. The large

  5. Lateral Gene Transfer and Gene Duplication Played a Key Role in the Evolution of Mastigamoeba balamuthi Hydrogenosomes

    PubMed Central

    Nývltová, Eva; Stairs, Courtney W.; Hrdý, Ivan; Rídl, Jakub; Mach, Jan; Pačes, Jan; Roger, Andrew J.; Tachezy, Jan

    2015-01-01

    Lateral gene transfer (LGT) is an important mechanism of evolution for protists adapting to oxygen-poor environments. Specifically, modifications of energy metabolism in anaerobic forms of mitochondria (e.g., hydrogenosomes) are likely to have been associated with gene transfer from prokaryotes. An interesting question is whether the products of transferred genes were directly targeted into the ancestral organelle or initially operated in the cytosol and subsequently acquired organelle-targeting sequences. Here, we identified key enzymes of hydrogenosomal metabolism in the free-living anaerobic amoebozoan Mastigamoeba balamuthi and analyzed their cellular localizations, enzymatic activities, and evolutionary histories. Additionally, we characterized 1) several canonical mitochondrial components including respiratory complex II and the glycine cleavage system, 2) enzymes associated with anaerobic energy metabolism, including an unusual D-lactate dehydrogenase and acetyl CoA synthase, and 3) a sulfate activation pathway. Intriguingly, components of anaerobic energy metabolism are present in at least two gene copies. For each component, one copy possesses an mitochondrial targeting sequence (MTS), whereas the other lacks an MTS, yielding parallel cytosolic and hydrogenosomal extended glycolysis pathways. Experimentally, we confirmed that the organelle targeting of several proteins is fully dependent on the MTS. Phylogenetic analysis of all extended glycolysis components suggested that these components were acquired by LGT. We propose that the transformation from an ancestral organelle to a hydrogenosome in the M. balamuthi lineage involved the lateral acquisition of genes encoding extended glycolysis enzymes that initially operated in the cytosol and that established a parallel hydrogenosomal pathway after gene duplication and MTS acquisition. PMID:25573905

  6. Interleukin 10 protects primary melanocyte by activation of Stat-3 and PI3K/Akt/NF-κB signaling pathways.

    PubMed

    Zhou, Jia; Ling, Jingjing; Song, Jing; Wang, Yong; Feng, Bainian; Ping, Fengfeng

    2016-07-01

    Vitiligo is a common melanocytopenic disorder of the skin, with acquired focal depigmentation. Normal human skin relies on melanocytes to provide photoprotection and thermoregulation by producing melanin. Interleukin 10 (IL-10) is a pleiotropic immunoregulatory cytokine drawing more and more researchers' attention. The present study was conducted to investigate the effects of IL-10 on melanocytes and elucidate the underlying mechanisms. We proved that IL-10 play no role in regulating melanogenesis of normal human foreskin-derived epidermal melanocytes (NHEM). IL-10 stimulation activated the JAK/Stat-3 and PI3K/Akt signaling pathways. Moreover, IL-10 treatment increased translocation of p65 NF-κB into the nuclear compartment, and up-regulated expression of the pro-survival proteins Bcl-2 and Bcl-xL. IL-10 restored anti-apoptotic proteins expression and suppressed cytochrome c release in H2O2-induced apoptosis. In conclusion, IL-10 may provide pro-survival cues to melanocytes and be applied in the treatment of vitiligo and other depigmenting disorders. PMID:27186967

  7. Molecular characterization of woodchuck interleukin-10 receptor and enhanced function of specific T cells from chronically infected woodchucks following its blockade.

    PubMed

    Jiang, Min; Liu, Jia; Zhang, Ejuan; Meng, Zhongji; Wang, Baoju; Roggendorf, Michael; Yang, Dongliang; Lu, Mengji; Xu, Yang

    2012-12-01

    Interleukin 10 (IL-10) is a pleiotropic cytokine acting on a variety of immune cells through the cell surface receptor (IL-10R). It has been suggested to resuscitate antiviral immunity by interfering with IL-10/IL-10R pathway. The woodchuck model infected by woodchuck hepatitis virus (WHV) represents an informative animal model to study hepatitis B virus (HBV) infection. In this study, the woodchuck IL-10R (wIL-10R) was molecularly cloned and characterized, showing high similarity of its nucleotide and amino acid sequences to that of other mammalian species. The expression level of wIL-10R mRNA in woodchuck peripheral blood mononuclear cells was significantly increased in acute WHV infection but down-regulated during chronic WHV infection. Specific rabbit antibodies against wIL-10R were prepared and showed the ability to enhance the proliferation and degranulation of specific T-cells from chronically WHV-infected woodchucks in vitro. The present work on wIL-10R provided a good basis for future preclinical studies on therapeutic approaches for chronic HBV infection. PMID:22784930

  8. Interleukin 10 inhibits macrophage microbicidal activity by blocking the endogenous production of tumor necrosis factor alpha required as a costimulatory factor for interferon gamma-induced activation.

    PubMed Central

    Oswald, I P; Wynn, T A; Sher, A; James, S L

    1992-01-01

    Interleukin 10 (IL-10) inhibits interferon gamma-induced macrophage activation for cytotoxicity against larvae of the human parasite Schistosoma mansoni by suppressing production of the toxic effector molecule nitric oxide (NO). In this study, the mechanism of IL-10 action was identified as inhibition of endogenous tumor necrosis factor alpha (TNF-alpha) production by interferon gamma-activated macrophages. TNF-alpha appears to serve as a cofactor for interferon gamma-mediated activation, since both schistosomulum killing and NO production were inhibited by anti-TNF-alpha antibody, whereas TNF-alpha alone was unable to stimulate these macrophage functions. IL-10 blocked TNF-alpha production by interferon gamma-treated macrophages at the levels of both protein and mRNA synthesis. Addition of exogenous TNF-alpha reversed IL-10-mediated suppression of macrophage cytotoxic activity as well as NO production. Likewise, addition of a macrophage-triggering agent (bacterial lipopolysaccharide or muramyl dipeptide), which induced the production of TNF-alpha, also reversed the suppressive effect of IL-10 on cytotoxic function. In contrast to IL-10, two other cytokines, IL-4 and transforming growth factor beta, which also inhibit macrophage activation for schistosomulum killing and NO production, did not substantially suppress endogenous TNF-alpha production. These results, therefore, describe a separate pathway by which macrophage microbicidal function is inhibited by the down-regulatory cytokine IL-10. Images PMID:1528880

  9. Interleukin-10 antisense oligodeoxynucleotide suppresses IL-10 expression and effects on proinflammatory cytokine responses to porcine reproductive and respiratory syndrome virus.

    PubMed

    Charerntantanakul, Wasin; Kasinrerk, Watchara

    2010-08-01

    Upregulation of interleukin-10 (IL-10) expression has been suggested to be the mechanism by which the porcine reproductive and respiratory syndrome virus (PRRSV) suppresses the innate and adaptive immune response in infected pigs. In this study we evaluated the potential of phosphorothioate-modified IL-10 antisense oligodeoxynucleotide specific to the translation initiation region of porcine IL-10 mRNA (IL-10AS) in enhancing proinflammatory cytokine responses to PRRSV. Naïve peripheral blood mononuclear cells from eight PRRSV-seronegative pigs were transfected with IL-10AS in vitro prior to PRRSV inoculation and phorbol 12-myristate 13-acetate plus ionomycin or concanavalin A stimulation. The effects of IL-10AS on mRNA expression of IL-10, interferon-gamma (IFN-gamma), IFN-alpha, tumor necrosis factor-alpha (TNF-alpha), IL-2, and IL-4 were tested by real-time PCR. The percentages of IFN-gamma-producing T-cell subsets were determined by flow cytometry. Compared to the controls, the levels of IL-10 and IL-2 mRNA were significantly reduced, while those of IFN-gamma mRNA were increased, and TNF-alpha, IFN-alpha, and IL-4 mRNA were unchanged. An increase in the percentage of the IFN-gamma+ population was also observed in lymphocytes and CD8beta+ T cells. Our results suggest that IL-10AS has the potential to enhance proinflammatory cytokine responses to PRRSV infection. PMID:20712487

  10. Controlled dual delivery of fibroblast growth factor-2 and Interleukin-10 by heparin-based coacervate synergistically enhances ischemic heart repair.

    PubMed

    Chen, William C W; Lee, Brandon G; Park, Dae Woo; Kim, Kyobum; Chu, Hunghao; Kim, Kang; Huard, Johnny; Wang, Yadong

    2015-12-01

    Myocardial infarction (MI) causes myocardial necrosis, triggers chronic inflammatory responses, and leads to pathological remodeling. Controlled delivery of a combination of angiogenic and immunoregulatory proteins may be a promising therapeutic approach for MI. We investigated the bioactivity and therapeutic potential of an injectable, heparin-based coacervate co-delivering an angiogenic factor, fibroblast growth factor-2 (FGF2), and an anti-inflammatory cytokine, Interleukin-10 (IL-10) in a spatially and temporally controlled manner. Coacervate delivery of FGF2 and IL-10 preserved their bioactivities on cardiac stromal cell proliferation in vitro. Upon intramyocardial injection into a mouse MI model, echocardiography revealed that FGF2/IL-10 coacervate treated groups showed significantly improved long-term LV contractile function and ameliorated LV dilatation. FGF2/IL-10 coacervate substantially augmented LV myocardial elasticity. Additionally, FGF2/IL-10 coacervate notably enhanced long-term revascularization, especially at the infarct area. In addition, coacervate loaded with 500 ng FGF2 and 500 ng IL-10 significantly reduced LV fibrosis, considerably preserved infarct wall thickness, and markedly inhibited chronic inflammation at the infarct area. These results indicate that FGF2/IL-10 coacervate has notably greater therapeutic potential than coacervate containing only FGF2. Overall, our data suggest therapeutically synergistic effects of FGF-2/IL-10 coacervate, particularly coacervate with FGF2 and 500 ng IL-10, for the treatment of ischemic heart disease. PMID:26370927

  11. Comparative insights into the regulation of inflammation: Levels and predictors of interleukin 6 and interleukin 10 in young adults in the Philippines

    PubMed Central

    Tallman, Paula S.; Adair, Linda S.; Judith, Borja; Kuzawa, Christopher W.

    2013-01-01

    Inflammation is a central part of innate immunity, but its role in anti-pathogen defenses has been overshadowed by recent interest in the contribution of inflammation to a wide range of chronic degenerative diseases. Current research on chronic inflammation is conducted primarily in affluent populations with low levels of infectious disease; comparative research in different ecological settings is needed to advance understandings of the causes and consequences of variation in the regulation of inflammation. This paper investigates the levels and predictors of interleukin-6 (IL-6) and interleukin-10 (IL-10)–two cytokines important to the regulation of inflammation—in a large, population-based study in the Philippines. Concentrations of IL-6 and IL-10 were determined in N=1569 healthy young adults (20-22 yrs) in Metro Cebu, Philippines. IL-6 and IL-10 concentrations were positively correlated, and body mass index and symptoms of infectious disease were both associated with higher concentrations of IL-6 and IL-10. Median concentrations of IL-6 (1.0 pg/mL) and IL-10 (7.56 pg/mL) were substantially lower and higher, respectively, than levels reported for other populations based on a systematic review of prior research. This study contributes to a growing body of research in human ecological immunology, and suggests that there may be substantial population differences in the regulation of inflammation that has implications for the association between inflammation and disease. PMID:21994014

  12. Goat Milk Yoghurt by Using Lacto-B Culture Modulates the Production of Tumor Necrosis Factor-Alpha and Interleukin-10 in Malnourished Rats

    PubMed Central

    Kandarina, BJ. Istiti; Kusuma, Sari; Trisnasari, Yunita Dewi

    2014-01-01

    Total spleen lymphocytes, lymphocyte proliferation, tumor necrosis factor-α (TNF-α), and interleukin-10 (IL-10) in spleen lymphocyte culture were studied in malnourished Wistar rats fed with goat milk yoghurt. Malnourished rats were created by using standard feed restriction as much as 50% of normal rats for 21 d. Goat milk yoghurt containing three types of microorganism e.g., Lactobacillus acidophilus, Sterptococcus thermophilus and Bifidobacterium longum derived from Lacto-B culture in powder form. After 21 d, the rats continued to receive restricted feeding and supplemented with goat milk yoghurt for 7 d. Total splenocytes were counted by hemocytometer. Splenocytes proliferation was expressed as stimulation index, whereas the TNF-α and IL-10 of spleen lymphocyte culture were measured by ELISA technique. The total number of splenocytes and stimulation index of splenocytes in moderate malnourished and normal rats supplemented with goat milk yoghurt was not significantly different. The level of TNF-α in the rat supplemented with goat milk yoghurt was lower (p<0.05) than the control group, whereas the level of IL-10 in the rat supplemented with goat milk yoghurt was higher (p<0.05) than the control group. In conclusion, goat milk yoghurt supplementation in malnourished rats could decrease TNF-α as a representation of the proinflammatory cytokine, while it increases IL-10 as a representation of the anti-inflammatory cytokine. PMID:26760750

  13. A Glucuronoxylomannan-Associated Immune Signature, Characterized by Monocyte Deactivation and an Increased Interleukin 10 Level, Is a Predictor of Death in Cryptococcal Meningitis

    PubMed Central

    Scriven, James E.; Graham, Lisa M.; Schutz, Charlotte; Scriba, Thomas J.; Wilkinson, Katalin A.; Wilkinson, Robert J.; Boulware, David R.; Urban, Britta C.; Lalloo, David G.; Meintjes, Graeme

    2016-01-01

    Background. Cryptococcal meningitis remains a significant cause of death among human immunodeficiency virus type 1 (HIV)–infected persons in Africa. We aimed to better understand the pathogenesis and identify immune correlates of mortality, particularly the role of monocyte activation. Methods. A prospective cohort study was conducted in Cape Town, South Africa. Patients with a first episode of cryptococcal meningitis were enrolled, and their immune responses were assessed in unstimulated and stimulated blood specimens, using flow cytometry and cytokine analysis. Results. Sixty participants were enrolled (median CD4+ T-cell count, 34 cells/µL). Mortality was 23% (14 of 60 participants) at 14 days and 39% (22 of 57) at 12 weeks. Nonsurvivors were more likely to have an altered consciousness and higher cerebrospinal fluid fungal burden at presentation. Principal component analysis identified an immune signature associated with early mortality, characterized by monocyte deactivation (reduced HLA-DR expression and tumor necrosis factor α response to lipopolysaccharide); increased serum interleukin 6, CXCL10, and interleukin 10 levels; increased neutrophil counts; and decreased T-helper cell type 1 responses. This immune signature remained an independent predictor of early mortality after adjustment for consciousness level and fungal burden and was associated with higher serum titers of cryptococcal glucuronoxylomannan. Conclusions. Cryptococcal-related mortality is associated with monocyte deactivation and an antiinflammatory blood immune signature, possibly due to Cryptococcus modulation of the host immune response. Validation in other cohorts is required. PMID:26768248

  14. High levels of interleukin-10 impair resistance to pulmonary coccidioidomycosis in mice in part through control of nitric oxide synthase 2 expression.

    PubMed

    Jimenez, Maria del Pilar; Walls, Lorraine; Fierer, Joshua

    2006-06-01

    We have shown previously that there is a direct correlation between IL-10 levels and susceptibility to Coccidioides immitis peritonitis in C57BL/6 (B6), DBA/2, and BXD recombinant inbred mice. We now show that B6 mice are also more susceptible to C. immitis pneumonia and that interleukin-10 (IL-10)-deficient (IL-10-/-) B6 mice are more resistant to C. immitis pneumonia. In addition, we established that high levels of IL-10 are sufficient to make genetically resistant mice susceptible to both C. immitis peritonitis and pneumonia by infecting h.IL-10 transgenic mice. Infected h.IL-10 transgenic mice express lower levels of gamma interferon, IL-12 p40, and inducible nitric oxide synthetase 2 (NOS2) mRNA in their lungs, implicating inducible NOS as a defense mechanism in this disease. We treated DBA/2 mice with aminoguanidine, and they became more susceptible to C. immitis peritonitis and pneumonia. We conclude that high levels of IL-10 are both necessary and sufficient to make mice susceptible to C. immitis, regardless of the genetic background of the mice, and that IL-10 impairs resistance to C. immitis in part by suppressing NO synthesis. PMID:16714569

  15. Local immunosuppression induced by high viral load of human papillomavirus: characterization of cellular phenotypes producing interleukin-10 in cervical neoplastic lesions.

    PubMed

    Prata, Thiago Theodoro Martins; Bonin, Camila Mareti; Ferreira, Alda Maria Teixeira; Padovani, Cacilda Tezelli Junqueira; Fernandes, Carlos Eurico dos Santos; Machado, Ana Paula; Tozetti, Inês Aparecida

    2015-09-01

    A specific immune response to human papillomavirus (HPV) in the cervical microenvironment plays a key role in eradicating infection and eliminating mutated cells. However, high-risk HPVs modulate immune cells to create an immunosuppressive microenvironment, and induce these immune cells to produce interleukin 10 (IL-10). This production of IL-10, in conjunction with HPV infection, contributes to the appearance of cervical neoplastic lesions. We sought to characterize the IL-10-producing cellular phenotype, and investigate the influence of host and HPV factors upon the induction of an immunosuppressive microenvironment. Immunohistochemical analysis demonstrated an increase in IL-10 production by keratinocytes, macrophages and Langerhans cells in high-grade cervical lesions and cervical cancer. This increase was more pronounced in patients older than 30 years, and was also correlated with high viral load, and infection with a single HPV type, particularly high-risk HPVs. Our results indicate the existence of a highly immunosuppressive microenvironment composed of different IL-10-producing cellular phenotypes in cervical cancer samples, and samples classified as high-grade cervical lesions (cervical intraepithelial neoplasia stages II and III). The immunosuppressive microenvironment that developed for these different cellular phenotypes favours viral persistence and neoplastic progression. PMID:26059395

  16. Lateralised behaviour and immune response in dogs: relations between paw preference and interferon-gamma, interleukin-10 and IgG antibodies production.

    PubMed

    Quaranta, A; Siniscalchi, M; Frate, A; Iacoviello, R; Buonavoglia, C; Vallortigara, G

    2006-01-30

    The production of specific antibodies (IgG), interleukin-10 (IL-10) and interferon-gamma (IFN-gamma) was evaluated in dogs in relation to behavioural lateralisation as assessed by paw preference. Left-handed, right-handed and ambidextrous dogs of mixed breed were selected on the basis of their performance in a task consisting of the removal of a piece of adhesive paper from the snout. All dogs were immunised with rabies vaccine. IgG anti-rabies antibody response was evaluated by indirect immunofluorescence (IIF) test. Serum IFN-gamma and IL-10 levels were measured by ELISA in animals showing significant individual left-, right- or no-paw preferences in the behavioural test. The results showed that the direction of behavioural lateralisation influenced the immune response in dogs. The titers of anti-rabies antibodies were lower in left-pawed dogs than in right-pawed and ambidextrous dogs. Similarly, the IFN-gamma serum levels were lower in left-pawed dogs than in right-pawed and ambidextrous dogs. IL-10, on the contrary, seemed to be an immune parameter, which was not affected by lateralisation. These findings suggest that immunomodulation can be correlated with brain laterality in canine species by the regulation of the production of antibodies and some cytokines like IFN-gamma, which are molecules involved in the immune-neurohumoral crosstalk. PMID:16159674

  17. Computational Modeling Predicts Interleukin-10 Control of Lesion Sterilization By Balancing Early Host-Immunity-Mediated Antimicrobial Responses With Caseation During Mycobacterium tuberculosis Infection

    PubMed Central

    Cilfone, Nicholas A.; Ford, Christopher B.; Marino, Simeone; Mattila, Joshua T.; Gideon, Hannah P.; Flynn, JoAnne L.; Kirschner, Denise E.; Linderman, Jennifer J.

    2014-01-01

    Although almost a third of the world’s population is infected with the bacterial pathogen Mycobacterium tuberculosis (Mtb), our understanding of the functions of many immune factors involved in fighting infection is limited. Determining the role of the immunosuppressive cytokine interleukin-10 (IL-10) at the level of the granuloma has proven difficult due to lesional heterogeneity and the limitations of animal models. Here we take an in silico approach and, through a series of virtual experiments, we predict several novel roles for IL-10 in TB granulomas: (1) decreased levels of IL-10 lead to increased numbers of sterile lesions, but at the cost of early increased caseation, (2) small increases in early antimicrobial activity cause this increased lesion sterility, (3) IL-10 produced by activated macrophages is a major mediator of early antimicrobial activity and early host-induced caseation and (4) increasing levels of infected macrophage derived IL-10 promotes bacterial persistence by limiting the early antimicrobial response and preventing lesion sterilization. Our findings, currently only accessible using an in silico approach, suggest that IL-10 at the individual granuloma scale is a critical regulator of lesion outcome. These predictions suggest IL-10 related mechanisms that could be used as adjunctive therapies during TB. PMID:25512604

  18. Study of Lateral Gene Transfer in an Acid Mine Drainage Community Enabled by Comparative Genomics

    NASA Astrophysics Data System (ADS)

    Hugenholtz, P.; Croft, L.; Tyson, G. W.; Baker, B. J.; Detter, C.; Richardson, P. M.; Banfield, J. F.

    2002-12-01

    Lateral gene transfer (LGT) is thought to play a crucial role in the ecology and evolution of prokaryotes. We are investigating the role of LGT in an acid mine drainage community hosted in a pyrite-dominated metal sulfide deposit at the Richmond mine at Iron Mountain, CA. Due to biologically-mediated pyrite dissolution, the prevailing conditions within the mine are extremely low pH (< 1.0), very high ionic concentrations (molar concentrations of iron sulfate and mM concentrations of arsenic, copper and zinc), and moderate to high temperatures (30 to >50 C). These conditions are thought to largely isolate the community from potential external gene donors since naked DNA, phage and prokaryotes native to neutral pH habitats do not persist at pH <1.0 precluding an external influx of genes by transformation, transduction and conjugation, respectively. Microbial communities exist in several distinct habitats within Richmond mine including biofilms (subaqueous slime streamers and subaerial slimes) and cells attached directly to pyrite granules. This, however, belies an unusual simplicity in community composition. All communities investigated to date comprise only a handful of phylogenetically distinct organisms, typically dominated by the iron-oxidizing genera Leptospirillum and Ferroplasma. We have undertaken a community genomics analysis of a subaerial biofilm dominated by a Leptospirillum population to facilitate the study of LGT in this type of environment. The genome of Ferroplasma acidarmanus fer1, a minor component of the target community (but a major component of other Richmond mine communities), has been sequenced. Comparative genome analyses indicate that F. acidarmanus and the ancestor of two acidophilic Thermoplasma species belonging to the Euryarchaeota have traded many genes with phylogenetically remote acidophilic Sulfolobus species (Crenarchaeota). The putatively transferred sets of Sulfolobus genes in Ferroplasma and the Thermoplasma ancestor are distinct

  19. Characterization of DNA-hyaluronan matrix for sustained gene transfer.

    PubMed

    Kim, Angela; Checkla, Daniel M; Dehazya, Philip; Chen, Weiliam

    2003-06-01

    DNA-Hyaluronan (DNA-HA) matrix formulations intended for use as gene delivery systems have been developed and their potential for delivering DNA encoding a model therapeutic cytokine, platelet-derived growth factor (PDGF), has been evaluated. The results of enzyme-mediated release kinetics studies suggested that the rate of DNA release from the DNA-HA matrices could be modulated by changing the DNA loading or the degree of crosslinking. SEM imaging of the DNA-HA matrix showed that it was gradually eroded by enzymatic action. The results of gel electrophoresis suggested that there was some degree of interaction between DNA and native HA and that portions of the DNA released from the DNA-HA matrices were associated with crosslinked HA fragments. Only fractions of the DNA released from the DNA-HA matrices were free and the rest was entrapped by HA fragments, which could serve as a mechanism for DNA protection. The results from cell transfection studies using DNA samples collected during the course of release studies confirmed this hypothesis. The PDGF produced by transfection of the DNA released from DNA-HA matrices induced human dermal fibroblast cells to proliferate. PMID:12767709

  20. Identification of a Divided Genome for VSH-1, the Prophage-Like Gene Transfer Agent of Brachyspira hyodysenteriae

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Brachyspira hyodysenteriae B204 genome sequence revealed three VSH-1 tail genes hvp31, hvp60, and hvp37, in a 3.6 kb cluster. The location and transcription direction of these genes relative to the previously described VSH-1 16.3 kb gene operon indicate that the gene transfer agent VSH-1 has a ...

  1. A case of horizontal gene transfer from Wolbachia to Aedes albopictus C6/36 cell line

    PubMed Central

    Hou, Qing; He, Ji; Yu, Jing; Ye, Yuting; Zhou, Dan; Sun, Yan; Zhang, Donghui; Ma, Lei; Shen, Bo; Zhu, Changliang

    2014-01-01

    Horizontal gene transfer plays an essential role in evolution and ecological adaptation, yet this phenomenon has remained controversial, particularly where it occurs between prokaryotes and eukaryotes. There are a handful of reported examples of horizontal gene transfer occurring between prokaryotes and eukaryotes in the literature, with most of these documented cases pertaining to invertebrates and endosymbionts. However, the vast majority of these horizontally transferred genes were either eventually excluded or rapidly became nonfunctional in the recipient genome. In this study, we report the discovery of a horizontal gene transfer from the endosymbiont Wolbachia in the C6/36 cell line derived from the mosquito Aedes albopictus. Moreover, we report that this horizontally transferred gene displayed high transcription level. This finding and the results of further experimentation strongly suggest this gene is functional and has been expressed and translated into a protein in the mosquito host cells. PMID:24812591

  2. Microbubbles and ultrasound increase intraventricular polyplex gene transfer to the brain.

    PubMed

    Tan, James-Kevin Y; Pham, Binhan; Zong, Yujin; Perez, Camilo; Maris, Don O; Hemphill, Ashton; Miao, Carol H; Matula, Thomas J; Mourad, Pierre D; Wei, Hua; Sellers, Drew L; Horner, Philip J; Pun, Suzie H

    2016-06-10

    Neurons in the brain can be damaged or lost from neurodegenerative disease, stroke, or traumatic injury. Although neurogenesis occurs in mammalian adult brains, the levels of natural neurogenesis are insufficient to restore function in these cases. Gene therapy has been pursued as a promising strategy to induce differentiation of neural progenitor cells into functional neurons. Non-viral vectors are a preferred method of gene transfer due to potential safety and manufacturing benefits but suffer from lower delivery efficiencies compared to viral vectors. Since the neural stem and progenitor cells reside in the subventricular zone of the brain, intraventricular injection has been used as an administration route for gene transfer to these cells. However, the choroid plexus epithelium remains an obstacle to delivery. Recently, transient disruption of the blood-brain barrier by microbubble-enhanced ultrasound has been used to successfully improve drug delivery to the brain after intravenous injection. In this work, we demonstrate that microbubble-enhanced ultrasound can similarly improve gene transfer to the subventricular zone after intraventricular injection. Microbubbles of different surface charges (neutral, slightly cationic, and cationic) were prepared, characterized by acoustic flow cytometry, and evaluated for their ability to increase the permeability of immortalized choroid plexus epithelium monolayers in vitro. Based on these results, slightly cationic microbubbles were evaluated for microbubble and ultrasound-mediated enhancement of non-viral gene transfer in vivo. When coupled with our previously reported gene delivery vehicles, the slightly cationic microbubbles significantly increased ultrasound-mediated transfection of the murine brain when compared to commercially available Definity® microbubbles. Temporary disruption of the choroid plexus by microbubble-enhanced ultrasound is therefore a viable way of enhancing gene delivery to the brain and merits

  3. Editing T cell specificity towards leukemia by zinc-finger nucleases and lentiviral gene transfer

    PubMed Central

    Lombardo, Angelo; Magnani, Zulma; Liu, Pei-Qi; Reik, Andreas; Chu, Victoria; Paschon, David E.; Zhang, Lei; Kuball, Jurgen; Camisa, Barbara; Bondanza, Attilio; Casorati, Giulia; Ponzoni, Maurilio; Ciceri, Fabio; Bordignon, Claudio; Greenberg, Philip D.; Holmes, Michael C.; Gregory, Philip D.; Naldini, Luigi; Bonini, Chiara

    2016-01-01

    The transfer of high-avidity T-cell receptor (TCR) genes isolated from rare tumor-specific lymphocytes into polyclonal T cells is an attractive cancer immunotherapy strategy. However, TCR gene transfer results in competition for surface expression and inappropriate pairing between the exogenous and endogenous TCR chains, resulting in suboptimal activity and potentially harmful unpredicted specificities. We designed zinc-finger nucleases (ZFNs) promoting the disruption of endogenous TCR β and α chain genes. ZFN-treated lymphocytes lacked CD3/TCR surface expression and expanded with IL-7 and IL-15. Upon lentiviral transfer of a TCR for the WT1 tumor antigen, these TCR-edited cells expressed the new TCR at high levels, were easily expanded to near-purity, and proved superior in specific antigen recognition to matched TCR-transferred cells. In contrast to TCR-transferred cells, TCR edited lymphocytes did not mediate off-target reactivity while maintaining anti-tumor activity in vivo, thus demonstrating that complete editing of T-cell specificity generate tumor-specific lymphocytes with improved biosafety profile. PMID:22466705

  4. Gene transfer rate from CL rice to diverse red rice biotypes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The future and sustainability of ClearfieldTM (CL) technology at the producers’ level will be dictated by various factors. Among many, prominent factors which affect the transfer of ALS-resistant gene from CL rice to red rice are: disparity in red rice biotypes and CL cultivars; flowering time of re...

  5. Bacteriophage-like Particles Associated with the Gene Transfer Agent of Methanococcus Voltale PS

    NASA Technical Reports Server (NTRS)

    Bertani, G.; Eiserling, F.; Pushkin, A.; Gingery, M.

    1999-01-01

    The methanogenic archaebacterium Methanococus voltae (strain PS) is known to produce a filterable, DNase resistant agent (called VTA, for voltae transfer agent), which carries very small fragments (4,400 base pairs) of bacterial DNA and is able to transduce bacterial genes between derivatives of the strain.

  6. Transfer of herbicide-resistant gene to weedy rice populations and its implications

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Red rice diversity in terms of phenology, sexual compatibility with cultivated rice, and the wide window of rice planting time can affect the rate of herbicide-resistant gene transfer from rice to RR. Experiments were conducted to a) determine the effect of red rice, rice cultivar, and planting date...

  7. Assessing the effects of a sequestered germline on interdomain lateral gene transfer in Metazoa.

    PubMed

    Jensen, Lindy; Grant, Jessica R; Laughinghouse, Haywood Dail; Katz, Laura A

    2016-06-01

    A sequestered germline in Metazoa has been argued to be an obstacle to lateral gene transfer (LGT), though few studies have specifically assessed this claim. Here, we test the hypothesis that the origin of a sequestered germline reduced LGT events in Bilateria (i.e., triploblast lineages) as compared to early-diverging Metazoa (i.e., Ctenophora, Cnidaria, Porifera, and Placozoa). We analyze single-gene phylogenies generated with over 900 species sampled from among Bacteria, Archaea, and Eukaryota to identify well-supported interdomain LGTs. We focus on ancient interdomain LGT (i.e., those between prokaryotes and multiple lineages of Metazoa) as systematic errors in single-gene tree reconstruction create uncertainties for interpreting eukaryote-to-eukaryote transfer. The breadth of the sampled Metazoa enables us to estimate the timing of LGTs, and to examine the pattern before versus after the evolution of a sequestered germline. We identified 58 LGTs found only in Metazoa and prokaryotes (i.e., bacteria and/or archaea), and seven genes transferred from prokaryotes into Metazoa plus one other eukaryotic clade. Our analyses indicate that more interdomain transfers occurred before the development of a sequestered germline, consistent with the hypothesis that this feature is an obstacle to LGT. PMID:27139503

  8. Diversity, evolution, and horizontal gene transfer (HGT) in soda lakes

    NASA Astrophysics Data System (ADS)

    Pinkart, Holly C.; Storrie-Lombardi, Michael C.

    2007-09-01

    Soap Lake is a hypersaline, alkaline lake in Central Washington State (USA). For the past five years the lake has been the site of an NSF Microbial Observatory project devoted to identifying critical geochemical and microbial characteristics of the monimolimnion sediment and water column, and has demonstrated rich multispecies communities occupy all areas of the lake. Soap Lake and similar soda lakes are subject to repeated transient periods of extreme evaporation characterized by significant repetitive alterations in salinity, pH, and total water volume, yet maintain high genetic and metabolic diversity. It has been argued that this repetitive cycle for salinity, alkalinity, and sulfur concentration has been a major driver for prokaryote evolution and diversity. The rapidity of wet-dry cycling places special demands on genome evolution, requirements that are beyond the relatively conservative eukaryotic evolutionary strategy of serial alteration of existing gene sequences in a relatively stable genome. Although HGT is most likely responsible for adding a significant amount of noise to the genetic record, analysis of HGT activity can also provide us with a much-needed probe for exploration of prokaryotic genome evolution and the origin of diversity. Packaging of genetic information within the protective protein capsid of a bacteriophage would seem preferable to exposing naked DNA to the highly alkaline conditions in the lake. In this study, we present preliminary data demonstrating the presence of a diverse group of phage integrases in Soap Lake. Integrase is the viral enzyme responsible for the insertion of phage DNA into the bacterial host's chromosome. The presence of the integrase sequence in bacterial chromosomes is evidence of lysogeny, and the diversity of integrase sequences reported here suggests a wide variety of temperate phage exist in this system, and are especially active in transition zones.

  9. Plasmid encoded antibiotic resistance: acquisition and transfer of antibiotic resistance genes in bacteria

    PubMed Central

    Bennett, P M

    2008-01-01

    Bacteria have existed on Earth for three billion years or so and have become adept at protecting themselves against toxic chemicals. Antibiotics have been in clinical use for a little more than 6 decades. That antibiotic resistance is now a major clinical problem all over the world attests to the success and speed of bacterial adaptation. Mechanisms of antibiotic resistance in bacteria are varied and include target protection, target substitution, antibiotic detoxification and block of intracellular antibiotic accumulation. Acquisition of genes needed to elaborate the various mechanisms is greatly aided by a variety of promiscuous gene transfer systems, such as bacterial conjugative plasmids, transposable elements and integron systems, that move genes from one DNA system to another and from one bacterial cell to another, not necessarily one related to the gene donor. Bacterial plasmids serve as the scaffold on which are assembled arrays of antibiotic resistance genes, by transposition (transposable elements and ISCR mediated transposition) and site-specific recombination mechanisms (integron gene cassettes). The evidence suggests that antibiotic resistance genes in human bacterial pathogens originate from a multitude of bacterial sources, indicating that the genomes of all bacteria can be considered as a single global gene pool into which most, if not all, bacteria can dip for genes necessary for survival. In terms of antibiotic resistance, plasmids serve a central role, as the vehicles for resistance gene capture and their subsequent dissemination. These various aspects of bacterial resistance to antibiotics will be explored in this presentation. PMID:18193080

  10. Plasmid encoded antibiotic resistance: acquisition and transfer of antibiotic resistance genes in bacteria.

    PubMed

    Bennett, P M

    2008-03-01

    Bacteria have existed on Earth for three billion years or so and have become adept at protecting themselves against toxic chemicals. Antibiotics have been in clinical use for a little more than 6 decades. That antibiotic resistance is now a major clinical problem all over the world attests to the success and speed of bacterial adaptation. Mechanisms of antibiotic resistance in bacteria are varied and include target protection, target substitution, antibiotic detoxification and block of intracellular antibiotic accumulation. Acquisition of genes needed to elaborate the various mechanisms is greatly aided by a variety of promiscuous gene transfer systems, such as bacterial conjugative plasmids, transposable elements and integron systems, that move genes from one DNA system to another and from one bacterial cell to another, not necessarily one related to the gene donor. Bacterial plasmids serve as the scaffold on which are assembled arrays of antibiotic resistance genes, by transposition (transposable elements and ISCR mediated transposition) and site-specific recombination mechanisms (integron gene cassettes).The evidence suggests that antibiotic resistance genes in human bacterial pathogens originate from a multitude of bacterial sources, indicating that the genomes of all bacteria can be considered as a single global gene pool into which most, if not all, bacteria can dip for genes necessary for survival. In terms of antibiotic resistance, plasmids serve a central role, as the vehicles for resistance gene capture and their subsequent dissemination. These various aspects of bacterial resistance to antibiotics will be explored in this presentation. PMID:18193080

  11. Ancient gene transfer from algae to animals: Mechanisms and evolutionary significance

    PubMed Central

    2012-01-01

    Background Horizontal gene transfer (HGT) is traditionally considered to be rare in multicellular eukaryotes such as animals. Recently, many genes of miscellaneous algal origins were discovered in choanoflagellates. Considering that choanoflagellates are the existing closest relatives of animals, we speculated that ancient HGT might have occurred in the unicellular ancestor of animals and affected the long-term evolution of animals. Results Through genome screening, phylogenetic and domain analyses, we identified 14 gene families, including 92 genes, in the tunicate Ciona intestinalis that are likely derived from miscellaneous photosynthetic eukaryotes. Almost all of these gene families are distributed in diverse animals, suggesting that they were mostly acquired by the common ancestor of animals. Their miscellaneous origins also suggest that these genes are not derived from a particular algal endosymbiont. In addition, most genes identified in our analyses are functionally related to molecule transport, cellular regulation and methylation signaling, suggesting that the acquisition of these genes might have facilitated the intercellular communication in the ancestral animal. Conclusions Our findings provide additional evidence that algal genes in aplastidic eukaryotes are not exclusively derived from historical plastids and thus important for interpreting the evolution of eukaryotic photosynthesis. Most importantly, our data represent the first evidence that more anciently acquired genes might exist in animals and that ancient HGT events have played an important role in animal evolution. PMID:22690978

  12. Conjugative transposons: an unusual and diverse set of integrated gene transfer elements.

    PubMed Central

    Salyers, A A; Shoemaker, N B; Stevens, A M; Li, L Y

    1995-01-01

    Conjugative transposons are integrated DNA elements that excise themselves to form a covalently closed circular intermediate. This circular intermediate can either reintegrate in the same cell (intracellular transposition) or transfer by conjugation to a recipient and integrate into the recipient's genome (intercellular transposition). Conjugative transposons were first found in gram-positive cocci but are now known to be present in a variety of gram-positive and gram-negative bacteria also. Conjugative transposons have a surprisingly broad host range, and they probably contribute as much as plasmids to the spread of antibiotic resistance genes in some genera of disease-causing bacteria. Resistance genes need not be carried on the conjugative transposon to be transferred. Many conjugative transposons can mobilize coresident plasmids, and the Bacteroides conjugative transposons can even excise and mobilize unlinked integrated elements. The Bacteroides conjugative transposons are also unusual in that their transfer activities are regulated by tetracycline via a complex regulatory network. PMID:8531886

  13. Fat-to-glucose interconversion by hydrodynamic transfer of two glyoxylate cycle enzyme genes

    PubMed Central

    Cordero, P; Campion, J; Milagro, FI; Marzo, F; Martinez, JA

    2008-01-01

    The glyoxylate cycle, which is well characterized in higher plants and some microorganisms but not in vertebrates, is able to bypass the citric acid cycle to achieve fat-to-carbohydrate interconversion. In this context, the hydrodynamic transfer of two glyoxylate cycle enzymes, such as isocytrate lyase (ICL) and malate synthase (MS), could accomplish the shift of using fat for the synthesis of glucose. Therefore, 20 mice weighing 23.37 ± 0.96 g were hydrodinamically gene transferred by administering into the tail vein a bolus with ICL and MS. After 36 hours, body weight, plasma glucose, respiratory quotient and energy expenditure were measured. The respiratory quotient was increased by gene transfer, which suggests that a higher carbohydrate/lipid ratio is oxidized in such animals. This application could help, if adequate protocols are designed, to induce fat utilization for glucose synthesis, which might be eventually useful to reduce body fat depots in situations of obesity and diabetes. PMID:19077206

  14. Lentivirus-mediated gene transfer to the central nervous system: therapeutic and research applications.

    PubMed

    Wong, Liang-Fong; Goodhead, Lucy; Prat, Christine; Mitrophanous, Kyriacos A; Kingsman, Susan M; Mazarakis, Nicholas D

    2006-01-01

    The management of disorders of the nervous system remains a medical challenge. The key goals are to understand disease mechanisms, to validate therapeutic targets, and to develop new therapeutic strategies. Viral vector-mediated gene transfer can meet these goals and vectors based on lentiviruses have particularly useful features. Lentiviral vectors can deliver 8 kb of sequence, they mediate gene transfer into any neuronal cell type, expression and therapy are sustained, and normal cellular functions in vitro and in vivo are not compromised. After delivery into the nervous system they induce no significant immune responses, there are no unwanted side effects of the vectors per se to date, and manufacturing and safety testing for clinical applications are well advanced. There are now numerous examples of effective long-term treatment of animal models of neurological disorders, such as Parkinson's disease, Alzheimer's disease, Huntington's disease, motor neuron diseases, lysosomal storage diseases, and spinal injury, using a range of therapeutic genes expressed in lentiviral vectors. Significant issues remain in some areas of neural gene therapy including defining the optimum therapeutic gene(s), increasing the specificity of delivery, regulating expression of potentially toxic genes, and designing clinically relevant strategies. We discuss the applications of lentiviral vectors in therapy and research and highlight the essential features that will ensure their translation to the clinic in the near future. PMID:16409120

  15. Horizontal gene transfer: essentiality and evolvability in prokaryotes, and roles in evolutionary transitions.

    PubMed

    Koonin, Eugene V

    2016-01-01

    The wide spread of gene exchange and loss in the prokaryotic world has prompted the concept of 'lateral genomics' to the point of an outright denial of the relevance of phylogenetic trees for evolution. However, the pronounced coherence congruence of the topologies of numerous gene trees, particularly those for (nearly) universal genes, translates into the notion of a statistical tree of life (STOL), which reflects a central trend of vertical evolution. The STOL can be employed as a framework for reconstruction of the evolutionary processes in the prokaryotic world. Quantitatively, however, horizontal gene transfer (HGT) dominates microbial evolution, with the rate of gene gain and loss being comparable to the rate of point mutations and much greater than the duplication rate. Theoretical models of evolution suggest that HGT is essential for the survival of microbial populations that otherwise deteriorate due to the Muller's ratchet effect. Apparently, at least some bacteria and archaea evolved dedicated vehicles for gene transfer that evolved from selfish elements such as plasmids and viruses. Recent phylogenomic analyses suggest that episodes of massive HGT were pivotal for the emergence of major groups of organisms such as multiple archaeal phyla as well as eukaryotes. Similar analyses appear to indicate that, in addition to donating hundreds of genes to the emerging eukaryotic lineage, mitochondrial endosymbiosis severely curtailed HGT. These results shed new light on the routes of evolutionary transitions, but caution is due given the inherent uncertainty of deep phylogenies. PMID:27508073

  16. Gene transfer in the liver using recombinant adeno-associated virus

    PubMed Central

    Ahmed, Seemin Seher; Li, Jia; Godwin, Jonathan; Gao, Guangping; Zhong, Li

    2013-01-01

    Liver-directed gene transfer and gene therapy are rapidly gaining attention primarily because the liver is centrally involved in a variety of metabolic functions that are affected in various inherited disorders. Recombinant adeno-associated virus (rAAV) is a popular gene delivery vehicle for gene therapy and intravenous delivery of some rAAV serotypes results in very efficient transduction of the liver. rAAV-mediated and liver-directed gene transfer can help in creating somatic transgenic animals or disease models and studying the function of various genes and miRNAs. The liver is the target tissue for gene therapy of many inborn metabolic diseases and may also be exploited as a “bio-factory” for the production of coagulation factors, insulin and growth hormones and other non-hepatic proteins. Hence efficient delivery of transgenes and small RNAs to the liver by rAAV vectors has been of long-standing interest to research scientists and clinicians alike. PMID:23686826

  17. Horizontal gene transfer: essentiality and evolvability in prokaryotes, and roles in evolutionary transitions

    PubMed Central

    Koonin, Eugene V.

    2016-01-01

    The wide spread of gene exchange and loss in the prokaryotic world has prompted the concept of ‘lateral genomics’ to the point of an outright denial of the relevance of phylogenetic trees for evolution. However, the pronounced coherence congruence of the topologies of numerous gene trees, particularly those for (nearly) universal genes, translates into the notion of a statistical tree of life (STOL), which reflects a central trend of vertical evolution. The STOL can be employed as a framework for reconstruction of the evolutionary processes in the prokaryotic world. Quantitatively, however, horizontal gene transfer (HGT) dominates microbial evolution, with the rate of gene gain and loss being comparable to the rate of point mutations and much greater than the duplication rate. Theoretical models of evolution suggest that HGT is essential for the survival of microbial populations that otherwise deteriorate due to the Muller’s ratchet effect. Apparently, at least some bacteria and archaea evolved dedicated vehicles for gene transfer that evolved from selfish elements such as plasmids and viruses. Recent phylogenomic analyses suggest that episodes of massive HGT were pivotal for the emergence of major groups of organisms such as multiple archaeal phyla as well as eukaryotes. Similar analyses appear to indicate that, in addition to donating hundreds of genes to the emerging eukaryotic lineage, mitochondrial endosymbiosis severely curtailed HGT. These results shed new light on the routes of evolutionary transitions, but caution is due given the inherent uncertainty of deep phylogenies. PMID:27508073

  18. Increased in vitro and in vivo gene transfer by adenovirus vectors containing chimeric fiber proteins.

    PubMed Central

    Wickham, T J; Tzeng, E; Shears, L L; Roelvink, P W; Li, Y; Lee, G M; Brough, D E; Lizonova, A; Kovesdi, I

    1997-01-01

    Alteration of the natural tropism of adenovirus (Ad) will permit gene transfer into specific cell types and thereby greatly broaden the scope of target diseases that can be treated by using Ad. We have constructed two Ad vectors which contain modifications to the Ad fiber coat protein that redirect virus binding to either alpha(v) integrin [AdZ.F(RGD)] or heparan sulfate [AdZ.F(pK7)] cellular receptors. These vectors were constructed by a novel method involving E4 rescue of an E4-deficient Ad with a transfer vector containing both the E4 region and the modified fiber gene. AdZ.F(RGD) increased gene delivery to endothelial and smooth muscle cells expressing alpha(v) integrins. Likewise, AdZ.F(pK7) increased transduction 5- to 500-fold in multiple cell types lacking high levels of Ad fiber receptor, including macrophage, endothelial, smooth muscle, fibroblast, and T cells. In addition, AdZ.F(pK7) significantly increased gene transfer in vivo to vascular smooth muscle cells of the porcine iliac artery following balloon angioplasty. These vectors may therefore be useful in gene therapy for vascular restenosis or for targeting endothelial cells in tumors. Although binding to the fiber receptor still occurs with these vectors, they demonstrate the feasibility of tissue-specific receptor targeting in cells which express low levels of Ad fiber receptor. PMID:9343173

  19. Intensive Pharmacological Immunosuppression Allows for Repetitive Liver Gene Transfer With Recombinant Adenovirus in Nonhuman Primates

    PubMed Central

    Fontanellas, Antonio; Hervás-Stubbs, Sandra; Mauleón, Itsaso; Dubrot, Juan; Mancheño, Uxua; Collantes, María; Sampedro, Ana; Unzu, Carmen; Alfaro, Carlos; Palazón, Asis; Smerdou, Cristian; Benito, Alberto; Prieto, Jesús; Peñuelas, Iván; Melero, Ignacio

    2010-01-01

    Repeated administration of gene therapies is hampered by host immunity toward vectors and transgenes. Attempts to circumvent antivector immunity include pharmacological immunosuppression or alternating different vectors and vector serotypes with the same transgene. Our studies show that B-cell depletion with anti-CD20 monoclonal antibody and concomitant T-cell inhibition with clinically available drugs permits repeated liver gene transfer to a limited number of nonhuman primates with recombinant adenovirus. Adenoviral vector–mediated transfer of the herpes simplex virus type 1 thymidine kinase (HSV1-tk) reporter gene was visualized in vivo with a semiquantitative transgene-specific positron emission tomography (PET) technique, liver immunohistochemistry, and immunoblot for the reporter transgene in needle biopsies. Neutralizing antibody and T cell–mediated responses toward the viral capsids were sequentially monitored and found to be repressed by the drug combinations tested. Repeated liver transfer of the HSV1-tk reporter gene with the same recombinant adenoviral vector was achieved in macaques undergoing a clinically feasible immunosuppressive treatment that ablated humoral and cellular immune responses. This strategy allows measurable gene retransfer to the liver as late as 15 months following the first adenoviral exposure in a macaque, which has undergone a total of four treatments with the same adenoviral vector. PMID:20087317

  20. Direct gene transfer into human cultured cells facilitated by laser micropuncture of the cell membrane

    SciTech Connect

    Tao, W.; Wilkinson, J.; Stanbridge, E.J.; Berns, M.W.

    1987-06-01

    The selective alteration of the cellular genome by laser microbeam irradiation has been extensively applied in cell biology. The authors report here the use of the third harmonic (355 nm) of an yttrium-aluminum garnet laser to facilitate the direct transfer of the neo gene into cultured human HT1080-6TG cells. The resultant transformants were selected in medium containing an aminoglycoside antibiotic, G418. Integration of the neo gene into individual human chromosomes and expression of the gene were demonstrated by Southern blot analyses, microcell-mediated chromosome transfer, and chromosome analyses. The stability of the integrated neo gene in the transformants was shown by a comparative growth assay in selective and nonselective media. Transformation and incorporation of the neo gene into the host genome occurred at a frequency of 8 x 10 /sup -4/-3 x 10/sup -3/. This method appears to be 100-fold more efficient than the standard calcium phosphate-mediated method of DNA transfer.

  1. Myocardial Gene Transfer: Routes and Devices for Regulation of Transgene Expression by Modulation of Cellular Permeability

    PubMed Central

    Katz, Michael G.; Bridges, Charles R.

    2013-01-01

    Abstract Heart diseases are major causes of morbidity and mortality in Western society. Gene therapy approaches are becoming promising therapeutic modalities to improve underlying molecular processes affecting failing cardiomyocytes. Numerous cardiac clinical gene therapy trials have yet to demonstrate strong positive results and advantages over current pharmacotherapy. The success of gene therapy depends largely on the creation of a reliable and efficient delivery method. The establishment of such a system is determined by its ability to overcome the existing biological barriers, including cellular uptake and intracellular trafficking as well as modulation of cellular permeability. In this article, we describe a variety of physical and mechanical methods, based on the transient disruption of the cell membrane, which are applied in nonviral gene transfer. In addition, we focus on the use of different physiological techniques and devices and pharmacological agents to enhance endothelial permeability. Development of these methods will undoubtedly help solve major problems facing gene therapy. PMID:23427834

  2. Source–sink plasmid transfer dynamics maintain gene mobility in soil bacterial communities

    PubMed Central

    Wood, A. Jamie

    2016-01-01

    Horizontal gene transfer is a fundamental process in bacterial evolution that can accelerate adaptation via the sharing of genes between lineages. Conjugative plasmids are the principal genetic elements mediating the horizontal transfer of genes, both within and between bacterial species. In some species, plasmids are unstable and likely to be lost through purifying selection, but when alternative hosts are available, interspecific plasmid transfer could counteract this and maintain access to plasmid-borne genes. To investigate the evolutionary importance of alternative hosts to plasmid population dynamics in an ecologically relevant environment, we established simple soil microcosm communities comprising two species of common soil bacteria, Pseudomonas fluorescens and Pseudomonas putida, and a mercury resistance (HgR) plasmid, pQBR57, both with and without positive selection [i.e., addition of Hg(II)]. In single-species populations, plasmid stability varied between species: although pQBR57 survived both with and without positive selection in P. fluorescens, it was lost or replaced by nontransferable HgR captured to the chromosome in P. putida. A simple mathematical model suggests these differences were likely due to pQBR57’s lower intraspecific conjugation rate in P. putida. By contrast, in two-species communities, both models and experiments show that interspecific conjugation from P. fluorescens allowed pQBR57 to persist in P. putida via source–sink transfer dynamics. Moreover, the replacement of pQBR57 by nontransferable chromosomal HgR in P. putida was slowed in coculture. Interspecific transfer allows plasmid survival in host species unable to sustain the plasmid in monoculture, promoting community-wide access to the plasmid-borne accessory gene pool and thus potentiating future evolvability. PMID:27385827

  3. Source-sink plasmid transfer dynamics maintain gene mobility in soil bacterial communities.

    PubMed

    Hall, James P J; Wood, A Jamie; Harrison, Ellie; Brockhurst, Michael A

    2016-07-19

    Horizontal gene transfer is a fundamental process in bacterial evolution that can accelerate adaptation via the sharing of genes between lineages. Conjugative plasmids are the principal genetic elements mediating the horizontal transfer of genes, both within and between bacterial species. In some species, plasmids are unstable and likely to be lost through purifying selection, but when alternative hosts are available, interspecific plasmid transfer could counteract this and maintain access to plasmid-borne genes. To investigate the evolutionary importance of alternative hosts to plasmid population dynamics in an ecologically relevant environment, we established simple soil microcosm communities comprising two species of common soil bacteria, Pseudomonas fluorescens and Pseudomonas putida, and a mercury resistance (Hg(R)) plasmid, pQBR57, both with and without positive selection [i.e., addition of Hg(II)]. In single-species populations, plasmid stability varied between species: although pQBR57 survived both with and without positive selection in P. fluorescens, it was lost or replaced by nontransferable Hg(R) captured to the chromosome in P. putida A simple mathematical model suggests these differences were likely due to pQBR57's lower intraspecific conjugation rate in P. putida By contrast, in two-species communities, both models and experiments show that interspecific conjugation from P. fluorescens allowed pQBR57 to persist in P. putida via source-sink transfer dynamics. Moreover, the replacement of pQBR57 by nontransferable chromosomal Hg(R) in P. putida was slowed in coculture. Interspecific transfer allows plasmid survival in host species unable to sustain the plasmid in monoculture, promoting community-wide access to the plasmid-borne accessory gene pool and thus potentiating future evolvability. PMID:27385827

  4. Targeting a newly established spontaneous feline fibrosarcoma cell line by gene transfer.

    PubMed

    Nande, Rounak; Di Benedetto, Altomare; Aimola, Pierpaolo; De Carlo, Flavia; Carper, Miranda; Claudio, Charlene D; Denvir, Jim; Valluri, Jagan; Duncan, Gary C; Claudio, Pier Paolo

    2012-01-01

    Fibrosarcoma is a deadly disease in cats and is significantly more often located at classical vaccine injections sites. More rare forms of spontaneous non-vaccination site (NSV) fibrosarcomas have been described and have been found associated to genetic alterations. Purpose of this study was to compare the efficacy of adenoviral gene transfer in NVS fibrosarcoma. We isolated and characterized a NVS fibrosarcoma cell line (Cocca-6A) from a spontaneous fibrosarcoma that occurred in a domestic calico cat. The feline cells were karyotyped and their chromosome number was counted using a Giemsa staining. Adenoviral gene transfer was verified by western blot analysis. Flow cytometry assay and Annexin-V were used to study cell-cycle changes and cell death of transduced cells. Cocca-6A fibrosarcoma cells were morphologically and cytogenetically characterized. Giemsa block staining of metaphase spreads of the Cocca-6A cells showed deletion of one of the E1 chromosomes, where feline p53 maps. Semi-quantitative PCR demonstrated reduction of p53 genomic DNA in the Cocca-6A cells. Adenoviral gene transfer determined a remarkable effect on the viability and growth of the Cocca-6A cells following single transduction with adenoviruses carrying Mda-7/IL-24 or IFN-γ or various combination of RB/p105, Ras-DN, IFN-γ, and Mda-7 gene transfer. Therapy for feline fibrosarcomas is often insufficient for long lasting tumor eradication. More gene transfer studies should be conducted in order to understand if these viral vectors could be applicable regardless the origin (spontaneous vs. vaccine induced) of feline fibrosarcomas. PMID:22666387

  5. Targeting a Newly Established Spontaneous Feline Fibrosarcoma Cell Line by Gene Transfer

    PubMed Central

    Nande, Rounak; De Carlo, Flavia; Carper, Miranda; Claudio, Charlene D.; Denvir, Jim; Valluri, Jagan; Duncan, Gary C.; Claudio, Pier Paolo

    2012-01-01

    Fibrosarcoma is a deadly disease in cats and is significantly more often located at classical vaccine injections sites. More rare forms of spontaneous non-vaccination site (NSV) fibrosarcomas have been described and have been found associated to genetic alterations. Purpose of this study was to compare the efficacy of adenoviral gene transfer in NVS fibrosarcoma. We isolated and characterized a NVS fibrosarcoma cell line (Cocca-6A) from a spontaneous fibrosarcoma that occurred in a domestic calico cat. The feline cells were karyotyped and their chromosome number was counted using a Giemsa staining. Adenoviral gene transfer was verified by western blot analysis. Flow cytometry assay and Annexin-V were used to study cell-cycle changes and cell death of transduced cells. Cocca-6A fibrosarcoma cells were morphologically and cytogenetically characterized. Giemsa block staining of metaphase spreads of the Cocca-6A cells showed deletion of one of the E1 chromosomes, where feline p53 maps. Semi-quantitative PCR demonstrated reduction of p53 genomic DNA in the Cocca-6A cells. Adenoviral gene transfer determined a remarkable effect on the viability and growth of the Cocca-6A cells following single transduction with adenoviruses carrying Mda-7/IL-24 or IFN-γ or various combination of RB/p105, Ras-DN, IFN-γ, and Mda-7 gene transfer. Therapy for feline fibrosarcomas is often insufficient for long lasting tumor eradication. More gene transfer studies should be conducted in order to understand if these viral vectors could be applicable regardless the origin (spontaneous vs. vaccine induced) of feline fibrosarcomas. PMID:22666387

  6. Networks of lexical borrowing and lateral gene transfer in language and genome evolution

    PubMed Central

    List, Johann-Mattis; Nelson-Sathi, Shijulal; Geisler, Hans; Martin, William

    2014-01-01

    Like biological species, languages change over time. As noted by Darwin, there are many parallels between language evolution and biological evolution. Insights into these parallels have also undergone change in the past 150 years. Just like genes, words change over time, and language evolution can be likened to genome evolution accordingly, but what kind of evolution? There are fundamental differences between eukaryotic and prokaryotic evolution. In the former, natural variation entails the gradual accumulation of minor mutations in alleles. In the latter, lateral gene transfer is an integral mechanism of natural variation. The study of language evolution using biological methods has attracted much interest of late, most approaches focusing on language tree construction. These approaches may underestimate the important role that borrowing plays in language evolution. Network approaches that were originally designed to study lateral gene transfer may provide more realistic insights into the complexities of language evolution. PMID:24375688

  7. Parallel Evolution and Horizontal Gene Transfer of the pst Operon in Firmicutes from Oligotrophic Environments

    PubMed Central

    Moreno-Letelier, Alejandra; Olmedo, Gabriela; Eguiarte, Luis E.; Martinez-Castilla, Leon; Souza, Valeria

    2011-01-01

    The high affinity phosphate transport system (pst) is crucial for phosphate uptake in oligotrophic environments. Cuatro Cienegas Basin (CCB) has extremely low P levels and its endemic Bacillus are closely related to oligotrophic marine Firmicutes. Thus, we expected the pst operon of CCB to share the same evolutionary history and protein similarity to marine Firmicutes. Orthologs of the pst operon were searched in 55 genomes of Firmicutes and 13 outgroups. Phylogenetic reconstructions were performed for the pst operon and 14 concatenated housekeeping genes using maximum likelihood methods. Conserved domains and 3D structures of the phosphate-binding protein (PstS) were also analyzed. The pst operon of Firmicutes shows two highly divergent clades with no correlation to the type of habitat nor a phylogenetic congruence, suggesting horizontal gene transfer. Despite sequence divergence, the PstS protein had a similar 3D structure, which could be due to parallel evolution after horizontal gene transfer events. PMID:21461370

  8. Parallel Evolution and Horizontal Gene Transfer of the pst Operon in Firmicutes from Oligotrophic Environments.

    PubMed

    Moreno-Letelier, Alejandra; Olmedo, Gabriela; Eguiarte, Luis E; Martinez-Castilla, Leon; Souza, Valeria

    2011-01-01

    The high affinity phosphate transport system (pst) is crucial for phosphate uptake in oligotrophic environments. Cuatro Cienegas Basin (CCB) has extremely low P levels and its endemic Bacillus are closely related to oligotrophic marine Firmicutes. Thus, we expected the pst operon of CCB to share the same evolutionary history and protein similarity to marine Firmicutes. Orthologs of the pst operon were searched in 55 genomes of Firmicutes and 13 outgroups. Phylogenetic reconstructions were performed for the pst operon and 14 concatenated housekeeping genes using maximum likelihood methods. Conserved domains and 3D structures of the phosphate-binding protein (PstS) were also analyzed. The pst operon of Firmicutes shows two highly divergent clades with no correlation to the type of habitat nor a phylogenetic congruence, suggesting horizontal gene transfer. Despite sequence divergence, the PstS protein had a similar 3D structure, which could be due to parallel evolution after horizontal gene transfer events. PMID:21461370

  9. Growth enhancement of shrimp (Litopenaeus schmitti) after transfer of tilapia growth hormone gene.

    PubMed

    Arenal, Amilcar; Pimentel, Rafael; Pimentel, Eulogio; Martín, Leonardo; Santiesteban, Dayamí; Franco, Ramón; Aleström, Peter

    2008-05-01

    Electroporation of Litopenaeus schmitti embryos was used to transfer the pE300tiGH15 plasmid that contains the tilapia growth hormone gene (tiGH) complexed with a nuclear localization signal peptide into the zygotes. The gene construct was detected in 35 (36%) of the 98 larvae screened by PCR and Southern blot analyses. Western blot analyses revealed that 34% of the screened larvae expressed a single tiGH-specific band with the expected molecular mass (23.1 kDa). The development index and larval length indicated a significant growth enhancement from day 3 on after electroporation, with an average of 32% of the growth enhancement. To our knowledge, this is the first report on gene transfer enhanced growth in crustaceans. PMID:18204820

  10. Detecting horizontal gene transfer with T-REX and RHOM programs.

    PubMed

    Li, Zuofeng; Wang, Li; Zhong, Yang

    2005-12-01

    As the Human Genome Project and other genome projects experience remarkable success and a flood of biological data is produced by means of high-throughout sequencing techniques, detection of horizontal gene transfer (HGT) becomes a promising field in bioinformatics. This review describes two freeware programs: T-REX for MS Windows and RHOM for Linux. T-REX is a graphical user interface program that offers functions to reconstruct the HGT network among the donor and receptor hosts from the gene and species distance matrices. RHOM is a set of command-line driven programs used to detect HGT in genomes. While T-REX impresses with a user-friendly interface and drawing of the reticulation network, the strength of RHOM is an extensive statistical framework of genome and the graphical display of the estimated sequence position probabilities for the candidate horizontally transferred genes. PMID:16420738

  11. Latest developments in gene transfer technology: achievements, perspectives, and controversies over therapeutic applications.

    PubMed

    Romano, G; Michell, P; Pacilio, C; Giordano, A

    2000-01-01

    Over the last decade, more than 300 phase I and phase II gene-based clinical trials have been conducted worldwide for the treatment of cancer and monogenic disorders. Lately, these trials have been extended to the treatment of AIDS and, to a lesser extent, cardiovascular diseases. There are 27 currently active gene therapy protocols for the treatment of HIV-1 infection in the USA. Preclinical studies are currently in progress to evaluate the possibility of increasing the number of gene therapy clinical trials for cardiopathies, and of beginning new gene therapy programs for neurologic illnesses, autoimmuno diseases, allergies, regeneration of tissues, and to implement procedures of allogeneic tissues or cell transplantation. In addition, gene transfer technology has allowed for the development of innovative vaccine design, known as genetic immunization. This technique has already been applied in the AIDS vaccine programs in the USA. These programs aim to confer protective immunity against HIV-1 transmission to individuals who are at risk of infection. Research programs have also been considered to develop therapeutic vaccines for patients with AIDS and generate either preventive or therapeutic vaccines against malaria, tuberculosis, hepatitis A, B and C viruses, influenza virus, La Crosse virus, and Ebola virus. The potential therapeutic applications of gene transfer technology are enormous. However, the effectiveness of gene therapy programs is still questioned. Furthermore, there is growing concern over the matter of safety of gene delivery and controversy has arisen over the proposal to begin in utero gene therapy clinical trials for the treatment of inherited genetic disorders. From this standpoint, despite the latest significant achievements reported in vector design, it is not possible to predict to what extent gene therapeutic interventions will be effective in patients, and in what time frame. PMID:10661569

  12. Effect of Plant and Environmental Factors on ALS-resistant Gene Transfer from ClearfieldTM Rice to Red Rice.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Imazethapyr-resistant gene from ClearfieldTM (CL) rice varieties transfers through pollen flow to red rice (Oryza sativa L.), a noxious weed in rice production in southern states. Factors which affect gene transfer rate include, but are not limited to, plant and environmental factors. Thus, we aimed...

  13. Intraspecies Transfer of the Chromosomal Acinetobacter baumannii blaNDM-1 Carbapenemase Gene.

    PubMed

    Krahn, Thomas; Wibberg, Daniel; Maus, Irena; Winkler, Anika; Bontron, Séverine; Sczyrba, Alexander; Nordmann, Patrice; Pühler, Alfred; Poirel, Laurent; Schlüter, Andreas

    2016-05-01

    The species Acinetobacter baumannii is one of the most important multidrug-resistant human pathogens. To determine its virulence and antibiotic resistance determinants, the genome of the nosocomial blaNDM-1-positive A. baumannii strain R2090 originating from Egypt was completely sequenced. Genome analysis revealed that strain R2090 is highly related to the community-acquired Australian A. baumannii strain D1279779. The two strains belong to sequence type 267 (ST267). Isolate R2090 harbored the chromosomally integrated transposon Tn125 carrying the carbapenemase gene blaNDM-1 that is not present in the D1279779 genome. To test the transferability of the metallo-β-lactamase (MBL) gene region, the clinical isolate R2090 was mated with the susceptible A. baumannii recipient CIP 70.10, and the carbapenem-resistant derivative R2091 was obtained. Genome sequencing of the R2091 derivative revealed that it had received an approximately 66-kb region comprising the transposon Tn125 embedding the blaNDM-1 gene. This region had integrated into the chromosome of the recipient strain CIP 70.10. From the four known mechanisms for horizontal gene transfer (conjugation, outer membrane vesicle-mediated transfer, transformation, and transduction), conjugation could be ruled out, since strain R2090 lacks any plasmid, and a type IV secretion system is not encoded in its chromosome. However, strain R2090 possesses three putative prophages, two of which were predicted to be complete and therefore functional. Accordingly, it was supposed that the transfer of the resistance gene region from the clinical isolate R2090 to the recipient occurred by general transduction facilitated by one of the prophages present in the R2090 genome. Hence, phage-mediated transduction has to be taken into account for the dissemination of antibiotic resistance genes within the species A. baumannii. PMID:26953198

  14. Immune Recognition of Gene Transfer Vectors: Focus on Adenovirus as a Paradigm

    PubMed Central

    Aldhamen, Yasser Ali; Seregin, Sergey S.; Amalfitano, Andrea

    2011-01-01

    Recombinant Adenovirus (Ad) based vectors have been utilized extensively as a gene transfer platform in multiple pre-clinical and clinical applications. These applications are numerous, and inclusive of both gene therapy and vaccine based approaches to human or animal diseases. The widespread utilization of these vectors in both animal models, as well as numerous human clinical trials (Ad-based vectors surpass all other gene transfer vectors relative to numbers of patients treated, as well as number of clinical trials overall), has shed light on how this virus vector interacts with both the innate and adaptive immune systems. The ability to generate and administer large amounts of this vector likely contributes not only to their ability to allow for highly efficient gene transfer, but also their elicitation of host immune responses to the vector and/or the transgene the vector expresses in vivo. These facts, coupled with utilization of several models that allow for full detection of these responses has predicted several observations made in human trials, an important point as lack of similar capabilities by other vector systems may prevent detection of such responses until only after human trials are initiated. Finally, induction of innate or adaptive immune responses by Ad vectors may be detrimental in one setting (i.e., gene therapy) and be entirely beneficial in another (i.e., prophylactic or therapeutic vaccine based applications). Herein, we review the current understanding of innate and adaptive immune responses to Ad vectors, as well some recent advances that attempt to capitalize on this understanding so as to further broaden the safe and efficient use of Ad-based gene transfer therapies in general. PMID:22566830

  15. Evolution and Horizontal Transfer of dUTPase-Encoding Genes in Viruses and Their Hosts

    PubMed Central

    Baldo, Angela M.; McClure, Marcella A.

    1999-01-01

    dUTPase is a ubiquitous and essential enzyme responsible for regulating cellular levels of dUTP. The dut gene exists as single, tandemly duplicated, and tandemly triplicated copies. Crystallized single-copy dUTPases have been shown to assemble as homotrimers. dUTPase is encoded as an auxiliary gene in a number of virus genomes. The origin of viral dut genes has remained unresolved since their initial discovery. A comprehensive analysis of dUTPase amino acid sequence relationships was performed to explore the evolutionary dynamics of dut in viruses and their hosts. Our data set, comprised of 24 host and 51 viral sequences, includes representative sequences from available eukaryotes, archaea, eubacteria cells, and viruses, including herpesviruses. These amino acid sequences were aligned by using a hidden Markov model approach developed to align divergent data. Known secondary structures from single-copy crystals were mapped onto the aligned duplicate and triplicate sequences. We show how duplicated dUTPases might fold into a monomer, and we hypothesize that triplicated dUTPases also assemble as monomers. Phylogenetic analysis revealed at least five viral dUTPase sequence lineages in well-supported monophyletic clusters with eukaryotic, eubacterial, and archaeal hosts. We have identified all five as strong examples of horizontal transfer as well as additional potential transfer of dut genes among eubacteria, between eubacteria and viruses, and between retroviruses. The evidence for horizontal transfers is particularly interesting since eukaryotic dut genes have introns, while DNA virus dut genes do not. This implies that an intermediary retroid agent facilitated the horizontal transfer process between host mRNA and DNA viruses. PMID:10438861

  16. Divergence of genes encoding non-specific lipid transfer proteins in the poaceae family.

    PubMed

    Jang, Cheol Seong; Jung, Jae Hyeong; Yim, Won Cheol; Lee, Byung-Moo; Seo, Yong Weon; Kim, Wook

    2007-10-31

    The genes encoding non-specific lipid transfer proteins (nsLTPs), members of a small multigene family, show a complex pattern of expressional regulation, suggesting that some diversification may have resulted from changes in their expression after duplication. In this study, the evolution of nsLTP genes within the Poaceae family was characterized via a survey of the pseudogenes and unigenes encoding the nsLTP in rice pseudomolecules and the NCBI unigene database. nsLTP-rich regions were detected in the distal portions of rice chromosomes 11 and 12; these may have resulted from the most recent large segmental duplication in the rice genome. Two independent tandem duplications were shown to occur within the nsLTP-rich regions of rice. The genomic distribution of the nsLTP genes in the rice genome differs from that in wheat. This may be attributed to gene migration, chromosomal rearrangement, and/or differential gene loss. The genomic distribution pattern of nsLTP genes in the Poaceae family points to the existence of some differences among cereal nsLTP genes, all of which diverged from an ancient gene. The unigenes encoding nsLTPs in each cereal species are clustered into five groups. The somewhat different distribution of nsLTP-encoding EST clones between the groups across cereal species imply that independent duplication(s) followed by subfunctionalization (and/or neofunctionalization) of the nsLTP gene family in each species occurred during speciation. PMID:17978574

  17. Evidence for extensive horizontal gene transfer from the draft genome of a tardigrade

    PubMed Central

    Boothby, Thomas C.; Tenlen, Jennifer R.; Smith, Frank W.; Wang, Jeremy R.; Patanella, Kiera A.; Osborne Nishimura, Erin; Tintori, Sophia C.; Li, Qing; Jones, Corbin D.; Yandell, Mark; Glasscock, Jarret; Goldstein, Bob

    2015-01-01

    Horizontal gene transfer (HGT), or the transfer of genes between species, has been recognized recently as more pervasive than previously suspected. Here, we report evidence for an unprecedented degree of HGT into an animal genome, based on a draft genome of a tardigrade, Hypsibius dujardini. Tardigrades are microscopic eight-legged animals that are famous for their ability to survive extreme conditions. Genome sequencing, direct confirmation of physical linkage, and phylogenetic analysis revealed that a large fraction of the H. dujardini genome is derived from diverse bacteria as well as plants, fungi, and Archaea. We estimate that approximately one-sixth of tardigrade genes entered by HGT, nearly double the fraction found in the most extreme cases of HGT into animals known to date. Foreign genes have supplemented, expanded, and even replaced some metazoan gene families within the tardigrade genome. Our results demonstrate that an unexpectedly large fraction of an animal genome can be derived from foreign sources. We speculate that animals that can survive extremes may be particularly prone to acquiring foreign genes. PMID:26598659

  18. Growth factor enhanced retroviral gene transfer to the adult central nervous system.

    PubMed

    King, L A; Mitrophanous, K A; Clark, L A; Kim, V N; Rohll, J B; Kingsman, A J; Colello, R J

    2000-07-01

    The use of viral vectors for gene delivery into mammalian cells provides a new approach in the treatment of many human diseases. The first viral vector approved for human clinical trials was murine leukemia virus (MLV), which remains the most commonly used vector in clinical trials to date. However, the application of MLV vectors is limited since MLV requires cells to be actively dividing in order for transduction and therefore gene delivery to occur. This limitation precludes the use of MLV for delivering genes to the adult CNS, where very little cell division is occurring. However, we speculated that this inherent limitation of ML V may be overcome by utilizing the known mitogenic effect of growth factors on cells of the CNS. Specifically, an in vivo application of growth factor to the adult brain, if able to induce cell division, could enhance MLV-based gene transfer to the adult brain. We now show that an exogenous application of basic fibroblast growth factor induces cell division in vivo. Under these conditions, where cells of the adult brain are stimulated to divide, MLV-based gene transfer is significantly enhanced. This novel approach precludes any vector modifications and provides a simple and effective way of delivering genes to cells of the adult brain utilizing MLV-based retroviral vectors. PMID:10918476

  19. Gene transfer of arginine kinase to skeletal muscle using adeno-associated virus

    PubMed Central

    Forbes, Sean C.; Bish, Lawrence T.; Ye, Fan; Spinazzola, Janelle; Baligand, Celine; Plant, Daniel; Vandenborne, Krista; Barton, Elisabeth R.; Sweeney, H. Lee; Walter, Glenn A.

    2014-01-01

    In this study we tested the feasibility of non-invasively measuring phosphoarginine (PArg) after gene delivery of arginine kinase (AK) using an adeno-associated virus (AAV) to murine hindlimbs. This was achieved by evaluating the time course, regional distribution, and metabolic flux of PArg using 31 phosphorus magnetic resonance spectroscopy (31P-MRS). AK gene was injected into the gastrocnemius of the left hindlimb of C57Bl10 mice (age 5wk, male) using self-complementary AAV, type 2/8 with desmin promoter. Non-localized 31P-MRS data were acquired over nine months after injection using 11.1-T and 17.6-T Bruker Avance spectrometers. In addition, 31P 2-D chemical shift imaging and saturation transfer experiments were performed to examine the spatial distribution and metabolic flux of PArg, respectively. PArg was evident in each injected mouse hindlimb after gene delivery, increased until 28 weeks, and remained elevated for at least nine months (p<.05). Furthermore, PArg was primarily localized to the injected posterior hindimb region with the metabolite being in exchange with ATP. Overall, the results show the viability of AAV gene transfer of AK gene to skeletal muscle, and provide support of PArg as a reporter that can be utilized to non-invasively monitor the transduction of genes for therapeutic interventions. PMID:24572791

  20. Targeted disruption of Ataxia-telangiectasia mutated gene in miniature pigs by somatic cell nuclear transfer

    SciTech Connect

    Kim, Young June; Ahn, Kwang Sung; Kim, Minjeong; Kim, Min Ju; Park, Sang-Min; Ryu, Junghyun; Ahn, Jin Seop; Heo, Soon Young; Kang, Jee Hyun; Choi, You Jung; Choi, Seong-Jun; Shim, Hosup

    2014-10-03

    Highlights: • ATM gene-targeted pigs were produced by somatic cell nuclear transfer. • A novel large animal model for ataxia telangiectasia was developed. • The new model may provide an alternative to the mouse model. - Abstract: Ataxia telangiectasia (A-T) is a recessive autosomal disorder associated with pleiotropic phenotypes, including progressive cerebellar degeneration, gonad atrophy, and growth retardation. Even though A-T is known to be caused by the mutations in the Ataxia telangiectasia mutated (ATM) gene, the correlation between abnormal cellular physiology caused by ATM mutations and the multiple symptoms of A-T disease has not been clearly determined. None of the existing ATM mouse models properly reflects the extent to which neurological degeneration occurs in human. In an attempt to provide a large animal model for A-T, we produced gene-targeted pigs with mutations in the ATM gene by somatic cell nuclear transfer. The disrupted allele in the ATM gene of cloned piglets was confirmed via PCR and Southern blot analysis. The ATM gene-targeted pigs generated in the present study may provide an alternative to the current mouse model for the study of mechanisms underlying A-T disorder and for the development of new therapies.

  1. Gene transfer of arginine kinase to skeletal muscle using adeno-associated virus.

    PubMed

    Forbes, S C; Bish, L T; Ye, F; Spinazzola, J; Baligand, C; Plant, D; Vandenborne, K; Barton, E R; Sweeney, H L; Walter, G A

    2014-04-01

    In this study, we tested the feasibility of non-invasively measuring phosphoarginine (PArg) after gene delivery of arginine kinase (AK) using an adeno-associated virus (AAV) to murine hindlimbs. This was achieved by evaluating the time course, regional distribution and metabolic flux of PArg using (31)phosphorus magnetic resonance spectroscopy ((31)P-MRS). AK gene was injected into the gastrocnemius of the left hindlimb of C57Bl10 mice (age 5 weeks, male) using self-complementary AAV, type 2/8 with desmin promoter. Non-localized (31)P-MRS data were acquired over 9 months after injection using 11.1-T and 17.6-T Bruker Avance spectrometers. In addition, (31)P two-dimensional chemical shift imaging and saturation transfer experiments were performed to examine the spatial distribution and metabolic flux of PArg, respectively. PArg was evident in each injected mouse hindlimb after gene delivery, increased until 28 weeks, and remained elevated for at least 9 months (P<0.05). Furthermore, PArg was primarily localized to the injected posterior hindimb region and the metabolite was in exchange with ATP. Overall, the results show the viability of AAV gene transfer of AK gene to skeletal muscle, and provide support of PArg as a reporter that can be used to non-invasively monitor the transduction of genes for therapeutic interventions. PMID:24572791

  2. Non-Invasive Gene Transfer by Iontophoresis for Therapy of an Inherited Retinal Degeneration

    PubMed Central

    Souied, Eric H.; Reid, Silvia N. M.; Piri, Natik I.; Lerner, Leonid E.; Nusinowitz, Steven; Farber, Debora B.

    2009-01-01

    Despite extensive research on many of the genes responsible for inherited retinal degenerations leading to blindness, no effective treatment is currently available for patients affected with these diseases. Among the therapeutic approaches tested on animal models of human retinal degeneration, gene therapy using different types of viral vectors as delivery agents has yielded promising results. We report here our results on a non-invasive, non-viral delivery approach using transscleral iontophoresis for transfer of plasmid DNA into mouse retina. Proof of principle experiments were carried out using plasmid containing GFP cDNA to demonstrate expression of the transferred gene in the retina after single applications of iontophoresis. Various parameters for multiple applications of iontophoresis were optimized to sustain GFP gene expression in mouse photoreceptors. Subsequently, repeated iontophoresis of plasmid containing normal β-phosphodiesterase (β-PDE) cDNA was performed in the rd1 mouse, an animal model of autosomal recessive retinitis pigmentosa caused by a mutant β-PDE gene. In normal mice, transscleral iontophoresis of the GFP plasmid provided a significant increase in fluorescence of the retina in the treated versus non-treated eyes. In rd1 mice, repeated iontophoresis of β-PDE cDNA plasmid partially rescued photoreceptors morphologically, as observed by microscopy, and functionally, as recorded on ERG measurements, without adverse effects. Therefore, transscleral iontophoresis of plasmid DNA containing therapeutic genes may be an efficient, safe and non-invasive method for the treatment of retinal degenerations. PMID:18653181

  3. Evidence for extensive horizontal gene transfer from the draft genome of a tardigrade.

    PubMed

    Boothby, Thomas C; Tenlen, Jennifer R; Smith, Frank W; Wang, Jeremy R; Patanella, Kiera A; Osborne Nishimura, Erin; Tintori, Sophia C; Li, Qing; Jones, Corbin D; Yandell, Mark; Messina, David N; Glasscock, Jarret; Goldstein, Bob

    2015-12-29

    Horizontal gene transfer (HGT), or the transfer of genes between species, has been recognized recently as more pervasive than previously suspected. Here, we report evidence for an unprecedented degree of HGT into an animal genome, based on a draft genome of a tardigrade, Hypsibius dujardini. Tardigrades are microscopic eight-legged animals that are famous for their ability to survive extreme conditions. Genome sequencing, direct confirmation of physical linkage, and phylogenetic analysis revealed that a large fraction of the H. dujardini genome is derived from diverse bacteria as well as plants, fungi, and Archaea. We estimate that approximately one-sixth of tardigrade genes entered by HGT, nearly double the fraction found in the most extreme cases of HGT into animals known to date. Foreign genes have supplemented, expanded, and even replaced some metazoan gene families within the tardigrade genome. Our results demonstrate that an unexpectedly large fraction of an animal genome can be derived from foreign sources. We speculate that animals that can survive extremes may be particularly prone to acquiring foreign genes. PMID:26598659

  4. An Adenovirus Vector Incorporating Carbohydrate Binding Domains Utilizes Glycans for Gene Transfer

    PubMed Central

    Nakayama, Masaharu; Ak, Ferhat; Ugai, Hideyo; Curiel, David T.

    2013-01-01

    Background Vectors based on human adenovirus serotype 5 (HAdV-5) continue to show promise as delivery vehicles for cancer gene therapy. Nevertheless, it has become clear that therapeutic benefit is directly linked to tumor-specific vector localization, highlighting the need for tumor-targeted gene delivery. Aberrant glycosylation of cell surface glycoproteins and glycolipids is a central feature of malignant transformation, and tumor-associated glycoforms are recognized as cancer biomarkers. On this basis, we hypothesized that cancer-specific cell-surface glycans could be the basis of a novel paradigm in HAdV-5-based vector targeting. Methodology/Principal Findings As a first step toward this goal, we constructed a novel HAdV-5 vector encoding a unique chimeric fiber protein that contains the tandem carbohydrate binding domains of the fiber protein of the NADC-1 strain of porcine adenovirus type 4 (PAdV-4). This glycan-targeted vector displays augmented CAR-independent gene transfer in cells with low CAR expression. Further, we show that gene transfer is markedly decreased in cells with genetic glycosylation defects and by inhibitors of glycosylation in normal cells. Conclusions/Significance These data provide the initial proof-of-concept for HAdV-5 vector-mediated gene delivery based on the presence of cell-surface carbohydrates. Further development of this new targeting paradigm could provide targeted gene delivery based on vector recognition of disease-specific glycan biomarkers. PMID:23383334

  5. Horizontal transfer of the msp130 gene supported the evolution of metazoan biomineralization.

    PubMed

    Ettensohn, Charles A

    2014-05-01

    It is widely accepted that biomineralized structures appeared independently in many metazoan clades during the Cambrian. How this occurred, and whether it involved the parallel co-option of a common set of biochemical and developmental pathways (i.e., a shared biomineralization "toolkit"), are questions that remain unanswered. Here, I provide evidence that horizontal gene transfer supported the evolution of biomineralization in some metazoans. I show that Msp130 proteins, first described as proteins expressed selectively by the biomineral-forming primary mesenchyme cells of the sea urchin embryo, have a much wider taxonomic distribution than was previously appreciated. Msp130 proteins are present in several invertebrate deuterostomes and in one protostome clade (molluscs). Surprisingly, closely related proteins are also present in many bacteria and several algae, and I propose that msp130 genes were introduced into metazoan lineages via multiple, independent horizontal gene transfer events. Phylogenetic analysis shows that the introduction of an ancestral msp130 gene occurred in the sea urchin lineage more than 250 million years ago and that msp130 genes underwent independent, parallel duplications in each of the metazoan phyla in which these genes are found. PMID:24735463

  6. Horizontal Gene Transfer Regulation in Bacteria as a “Spandrel” of DNA Repair Mechanisms

    PubMed Central

    Fall, Saliou; Mercier, Anne; Bertolla, Franck; Calteau, Alexandra; Gueguen, Laurent; Perrière, Guy; Vogel, Timothy M.; Simonet, Pascal

    2007-01-01

    Horizontal gene transfer (HGT) is recognized as the major force for bacterial genome evolution. Yet, numerous questions remain about the transferred genes, their function, quantity and frequency. The extent to which genetic transformation by exogenous DNA has occurred over evolutionary time was initially addressed by an in silico approach using the complete genome sequence of the Ralstonia solanacearum GMI1000 strain. Methods based on phylogenetic reconstruction of prokaryote homologous genes families detected 151 genes (13.3%) of foreign origin in the R. solanacearum genome and tentatively identified their bacterial origin. These putative transfers were analyzed in comparison to experimental transformation tests involving 18 different genomic DNA positions in the genome as sites for homologous or homeologous recombination. Significant transformation frequency differences were observed among these positions tested regardless of the overall genomic divergence of the R. solanacearum strains tested as recipients. The genomic positions containing the putative exogenous DNA were not systematically transformed at the highest frequencies. The two genomic “hot spots”, which contain recA and mutS genes, exhibited transformation frequencies from 2 to more than 4 orders of magnitude higher than positions associated with other genes depending on the recipient strain. These results support the notion that the bacterial cell is equipped with active mechanisms to modulate acquisition of new DNA in different genomic positions. Bio-informatics study correlated recombination “hot-spots” to the presence of Chi-like signature sequences with which recombination might be preferentially initiated. The fundamental role of HGT is certainly not limited to the critical impact that the very rare foreign genes acquired mainly by chance can have on the bacterial adaptation potential. The frequency to which HGT with homologous and homeologous DNA happens in the environment might have led

  7. Ionizing and ultraviolet radiation enhances the efficiency of DNA mediated gene transfer in vitro

    SciTech Connect

    Perez, C.F.

    1984-08-01

    The enhancement effects of ionizing and non-ionizing radiation on the efficiency of DNA mediated gene transfer were studied. Confluent Rat-2 cells were transfected with purified SV40 viral DNA, irradiated with either X-rays or ultraviolet, trypsinized, plated, and assayed for the formation of foci on Rat-2 monolayers. Both ionizing and ultraviolet radiation enhanced the frequency of A-gene transformants/survivor compared to unirradiated transfected cells. These enhancements were non-linear and dose dependent. A recombinant plasmid, pOT-TK5, was constructed that contained the SV40 virus A-gene and the Herpes Simplex virus (HSV) thymidine kinase (TK) gene. Confluent Rat-2 cells transfected with pOT-TK5 DNA and then immediately irradiated with either X-rays or 330 MeV/amu argon particles at the Berkeley Bevalac showed a higher frequency of HAT/sup +/ colonies/survivor than unirradiated transfected cells. Rat-2 cells transfected with the plasmid, pTK2, containing only the HSV TK-gene were enhanced for TK-transformation by both X-rays and ultraviolet radiation. The results demonstrate that radiation enhancement of the efficiency of DNA mediated gene transfer is not explained by increased nuclear uptake of the transfected DNA. Radiation increases the competence of the transfected cell population for genetic transformation. Three models for this increased competence are presented. The targeted integration model, the inducible recombination model, the partition model, and the utilization of DNA mediated gene transfer for DNA repair studies are discussed. 465 references.

  8. Hyperactive piggyBac Gene Transfer in Human Cells and In Vivo

    PubMed Central

    Doherty, Joseph E.; Huye, Leslie E.; Yusa, Kosuke; Zhou, Liqin; Craig, Nancy L.

    2012-01-01

    Abstract We characterized a recently developed hyperactive piggyBac (pB) transposase enzyme [containing seven mutations (7pB)] for gene transfer in human cells in vitro and to somatic cells in mice in vivo. Despite a protein level expression similar to that of native pB, 7pB significantly increased the gene transfer efficiency of a neomycin resistance cassette transposon in both HEK293 and HeLa cultured human cells. Native pB and SB100X, the most active transposase of the Sleeping Beauty transposon system, exhibited similar transposition efficiency in cultured human cell lines. When delivered to primary human T cells ex vivo, 7pB increased gene delivery two- to threefold compared with piggyBac and SB100X. The activity of hyperactive 7pB transposase was not affected by the addition of a 24-kDa N-terminal tag, whereas SB100X manifested a 50% reduction in transposition. Hyperactive 7pB was compared with native pB and SB100X in vivo in mice using hydrodynamic tail-vein injection of a limiting dose of transposase DNA combined with luciferase reporter transposons. We followed transgene expression for up to 6 months and observed approximately 10-fold greater long-term gene expression in mice injected with a codon-optimized version of 7pB compared with mice injected with native pB or SB100X. We conclude that hyperactive piggyBac elements can increase gene transfer in human cells and in vivo and should enable improved gene delivery using the piggyBac transposon system in a variety of cell and gene-therapy applications. PMID:21992617

  9. Oligosaccharide-specific induction of interleukin 10 production by B220+ cells from schistosome-infected mice: a mechanism for regulation of CD4+ T-cell subsets.

    PubMed Central

    Velupillai, P; Harn, D A

    1994-01-01

    Defining the factors and/or mechanisms that lead to the predominance of a particular CD4+ T-cell subset (Th-1 vs. Th-2) is an area of intense investigation. In murine schistosomiasis, Th-2-type T cells become predominant after deposition of eggs. The most immunoreactive egg components are glycoproteins. Previously we identified two interesting oligosaccharides found on schistosome eggs and schistosomula. One, lacto-N-fucopentaose III (LNFP-III) contains the interesting trisaccharide Lewisx, which is a weak ligand for P-selectin and is a sugar also found on the alpha and beta chains of the integrin lymphocyte function-associated molecule 1, a ligand for intercellular adhesion molecule 1. Because of the correlation between schistosome egg glycoproteins and Th-2 dominance, the present study examined whether LNFP-III and structurally related oligosaccharides were lymphostimulatory and/or able to induce factors known to down-regulate Th-1 cells. We found that LNFP-III and related sugars did induce proliferation of splenic non-T cells, B220+,CD4-,CD8- cells (B cells) of schistosome-infected and naive mice. In contrast to proliferation, LNFP-III was the only oligosaccharide that induced spleen cells to produce large amounts of interleukin 10 and prostaglandin E2, two molecules known to down-regulate Th-1 cells. Further, only spleen cells from infected mice produced cytokines after oligosaccharide stimulation. Interestingly, LNFP-III stimulation did not induce production of interleukin 4. Thus, a specific carbohydrate ligand has been identified that stimulates B cells to proliferate and produce factors that down-regulate Th-1 T cells. Further, we suggest that identical or structurally related ligands may contribute to the known Th-1 down-regulation in other parasitic diseases and in chronic blood-vascular diseases such as human immunodeficiency virus infection and a number of metastatic carcinomas and that this effect may, therefore, be a general phenomenon. PMID:7904066

  10. Genome-Wide Association Study Identifies That the ABO Blood Group System Influences Interleukin-10 Levels and the Risk of Clinical Events in Patients with Acute Coronary Syndrome

    PubMed Central

    Johansson, Åsa; Alfredsson, Jenny; Eriksson, Niclas; Wallentin, Lars; Siegbahn, Agneta

    2015-01-01

    Introduction Acute coronary syndrome (ACS) is a major cause of mortality worldwide. We have previously shown that increased interleukin-10 (IL-10) levels are associated with poor outcome in ACS patients. Method We performed a genome-wide association study in 2864 ACS patients and 408 healthy controls, to identify genetic variants associated with IL-10 levels. Then haplotype analyses of the identified loci were done and comparisons to levels of IL-10 and other known ACS related biomarkers. Results Genetic variants at the ABO blood group locus associated with IL-10 levels (top SNP: rs676457, P = 4.4 × 10−10) were identified in the ACS patients. Haplotype analysis, using SNPs tagging the four main ABO antigens (A1, A2, B and O), showed that O and A2 homozygous individuals, or O/A2 heterozygotes have much higher levels of IL-10 compared to individuals with other antigen combinations. In the ACS patients, associations between ABO antigens and von Willebrand factor (VWF, P = 9.2 × 10−13), and soluble tissue factor (sTF, P = 8.6 × 10−4) were also found. In the healthy control cohort, the associations with VWF and sTF were similar to those in ACS patients (P = 1.2 × 10−15 and P = 1.0 × 10−5 respectively), but the healthy cohort showed no association with IL-10 levels (P>0.05). In the ACS patients, the O antigen was also associated with an increased risk of cardiovascular death, all causes of death, and recurrent myocardial infarction (odds ratio [OR] = 1.24–1.29, P = 0.029–0.00067). Conclusion Our results suggest that the ABO antigens play important roles, not only for the immunological response in ACS patients, but also for the outcome of the disease. PMID:26600159

  11. Effect of scaling and root planing on serum interleukin-10 levels and glycemic control in chronic periodontitis and type 2 diabetes mellitus

    PubMed Central

    Acharya, Anirudh Balakrishna; Thakur, Srinath; Muddapur, Mahadevayya Veerayya

    2015-01-01

    Aim: Chronic periodontal disease (CPD) and type 2 diabetes mellitus (T2DM) share common pathogenic pathways involving the cytokine network resulting in increased susceptibility to both diseases, leading to increased inflammatory destruction, insulin resistance, and poor glycemic control. Periodontal treatment may improve glycemic control. The aim of this study was to evaluate the effect of scaling and root planing (SRP) of T2DM patients with CPD on hyperglycemia and the levels of serum interleukin-10 (IL-10). Materials and Methods: Forty-five subjects were divided into three groups comprising 15 subjects each as Group 1 (healthy controls), Group 2 (CPD patients), and Group 3 (T2DM patients with CPD). Plaque index, gingival index (GI), probing pocket depths (PPD), clinical attachment loss (AL), bleeding on probing (BoP), random blood sugar, glycosylated hemoglobin (HbA1C), and serum IL-10 were measured at baseline; SRP was performed on Groups 2 and 3 and the selected parameters recorded again at 6 months. Results: Statistically significant (P < 0.05) differences were observed in the variables at baseline and 6 months after SRP between the three groups using one-way ANOVA. The paired samples t-test for PPD and AL in Group 3 was statistically significant. Group 3 revealed positive correlations between PPD and HbA1C, BoP and IL-10, respectively, at 6 months and a predictable association of HbA1C with PPD and GI, and IL-10 levels with BoP, respectively, at 6 months. Conclusion: Scaling and root planing is effective in reducing blood glucose levels in T2DM patient with pocket depths and effective in elevating systemic IL-10 levels in CPD patients and CPD patients with T2DM. PMID:26015670

  12. Human Neutrophil Elastase Induce Interleukin-10 Expression in Peripheral Blood Mononuclear Cells through Protein Kinase C Theta/Delta and Phospholipase Pathways

    PubMed Central

    Kawata, Jin; Yamaguchi, Rui; Yamamoto, Takatoshi; Ishimaru, Yasuji; Sakamoto, Arisa; Aoki, Manabu; Kitano, Masafumi; Umehashi, Misako; Hirose, Eiji; Yamaguchi, Yasuo

    2016-01-01

    Objective Neutrophils have an important role in the rapid innate immune response, and the release or active secretion of elastase from neutrophils is linked to various inflammatory responses. Purpose of this study was to determine how the human neutrophil elastase affects the interleukin-10 (IL-10) response in peripheral blood mononuclear cells (PBMC). Materials and Methods In this prospective study, changes in IL-10 messenger RNA (mRNA) and protein expression levels in monocytes derived from human PBMCs were investigated after stimulation with human neutrophil elastase (HNE). A set of inhibitors was used for examining the pathways for IL-10 production induced by HNE. Results Reverse transcription polymerase chain reaction (RT-PCR) showed that stimulation with HNE upregulated IL-10 mRNA expression by monocytes, while the enzyme-linked immunosorbent assay (ELISA) revealed an increase of IL-10 protein level in the culture medium. A phospholipase C inhibitor (U73122) partially blunt- ed the induction of IL-10 mRNA expression by HNE, while IL-10 mRNA expression was significantly reduced by a protein kinase C (PKC) inhibitor (Rottlerin). A calcium chelator (3,4,5-trimethoxybenzoic acid 8-(diethylamino)octyl ester: TMB-8) inhibited the response of IL-10 mRNA to stimulation by HNE. In addition, pretreatment with a broad-spectrum PKC inhibitor (Ro-318425) partly blocked the response to HNE. Finally, an inhibitor of PKC theta/delta abolished the increased level of IL-10 mRNA expression. Conclusion These results indicate that HNE mainly upregulates IL-10 mRNA ex- pression and protein production in moncytes via a novel PKC theta/delta, although partially via the conventional PKC pathway. PMID:26862528

  13. Interleukin-10 conjugated electrospun polycaprolactone (PCL) nanofibre scaffolds for promoting alternatively activated (M2) macrophages around the peripheral nerve in vivo.

    PubMed

    Potas, Jason R; Haque, Farhia; Maclean, Francesca L; Nisbet, David R

    2015-05-01

    Macrophages play a key role in tissue regeneration following peripheral nerve injury by preparing the surrounding parenchyma for regeneration, however, they can be damaging if the response is excessive. Interleukin 10 (IL-10) is a cytokine that promotes macrophages toward an anti-inflammatory/wound healing state (M2 phenotype). The bioactive half-life of IL-10 is dependent on the cellular microenvironment and ranges from minutes to hours in vivo. Our objective was to extend the in vivo bioavailability and bioactivity of IL-10 by attaching the protein onto nanofibrous scaffolds and demonstrating increased expression levels of M2 macrophages when placed around healthy intact peripheral nerves. IL-10 was adsorbed and covalently bound to electrospun poly(ε-caprolactone) (PCL) nanofibrous scaffolds. In vivo bioavailability and bioactivity of IL-10 was confirmed by wrapping IL-10 conjugated nanofibres around the sciatic nerves of Wistar rats and quantifying M2 macrophages immunohistochemically double labelled with ED1 and either arginase 1 or CD206. IL-10 remained immobilised to PCL scaffolds for more than 120 days when stored in phosphate buffered saline at room temperature and for up to 14d ays when implanted around the sciatic nerve. IL-10 conjugated nanofibres successfully induced macrophage polarisation towards the M2 activated state within the scaffold material as well as the adjacent tissue surrounding the nerve. PCL biofunctionalised nanofibres are useful for manipulating the cellular microenvironment. Materials such as these could potentially lead to new therapeutic strategies for nervous tissue injuries as well as provide novel investigative tools for biological research. PMID:25837415

  14. Effects of amycenone on serum levels of tumor necrosis factor-α, interleukin-10, and depression-like behavior in mice after lipopolysaccharide administration.

    PubMed

    Yao, Wei; Zhang, Ji-chun; Dong, Chao; Zhuang, Cun; Hirota, Susumu; Inanaga, Kazutoyo; Hashimoto, Kenji

    2015-09-01

    Accumulating evidence suggests that inflammation plays a role in the pathophysiology of depression and that anti-inflammatory substances have antidepressant effects. Amycenone is obtained from extracts of the Yamabushitake (Hericium erinaceum). The purpose of this study is to examine whether amycenone shows anti-inflammatory and antidepressant effects in an inflammation-induced mouse model of depression. First, we examined the effects of amycenone on the serum levels of the pro-inflammatory cytokine, tumor necrosis factor-α (TNF-α), and the anti-inflammatory cytokine, interleukin-10 (IL-10), after intraperitoneal administration of the bacterial endotoxin lipopolysaccharide (LPS). Oral administration of amycenone (50, 100, or 200mg/kg) markedly blocked an increase in the serum TNF-α levels after a single administration of LPS (0.5mg/kg). Furthermore, amycenone (200mg/kg) markedly increased the serum IL-10 levels by a single administration of LPS (0.5mg/kg). Next, we examined the effects of amycenone on depression-like behaviors in the tail-suspension test (TST) and forced swimming test (FST). Pretreatment with amycenone (200mg/kg) significantly attenuated LPS (0.5mg/kg)-induced increase of the immobility time by the TST and FST, indicating antidepressant effects of amycenone. In addition, oral administration of paroxetine (30mg/kg) showed anti-inflammatory and antidepressant effects in the same model. These findings suggest that amycenone has antidepressant effects in LPS-induced inflammation model of depression. Therefore, amycenone could represent a potential supplement to prevent inflammation-related depression. PMID:26150007

  15. Multi-Scale Modeling Predicts a Balance of Tumor Necrosis Factor-α and Interleukin-10 Controls the Granuloma Environment during Mycobacterium tuberculosis Infection

    PubMed Central

    Cilfone, Nicholas A.; Perry, Cory R.; Kirschner, Denise E.; Linderman, Jennifer J.

    2013-01-01

    Interleukin-10 (IL-10) and tumor necrosis factor-α (TNF-α) are key anti- and pro-inflammatory mediators elicited during the host immune response to Mycobacterium tuberculosis (Mtb). Understanding the opposing effects of these mediators is difficult due to the complexity of processes acting across different spatial (molecular, cellular, and tissue) and temporal (seconds to years) scales. We take an in silico approach and use multi-scale agent based modeling of the immune response to Mtb, including molecular scale details for both TNF-α and IL-10. Our model predicts that IL-10 is necessary to modulate macrophage activation levels and to prevent host-induced tissue damage in a granuloma, an aggregate of cells that forms in response to Mtb. We show that TNF-α and IL-10 parameters related to synthesis, signaling, and spatial distribution processes control concentrations of TNF-α and IL-10 in a granuloma and determine infection outcome in the long-term. We devise an overall measure of granuloma function based on three metrics – total bacterial load, macrophage activation levels, and apoptosis of resting macrophages – and use this metric to demonstrate a balance of TNF-α and IL-10 concentrations is essential to Mtb infection control, within a single granuloma, with minimal host-induced tissue damage. Our findings suggest that a balance of TNF-α and IL-10 defines a granuloma environment that may be beneficial for both host and pathogen, but perturbing the balance could be used as a novel therapeutic strategy to modulate infection outcomes. PMID:23869227

  16. Delivery of interleukin-10 via injectable hydrogels improves renal outcomes and reduces systemic inflammation following ischemic acute kidney injury in mice.

    PubMed

    Soranno, Danielle E; Rodell, Christopher B; Altmann, Christopher; Duplantis, Jane; Andres-Hernando, Ana; Burdick, Jason A; Faubel, Sarah

    2016-08-01

    Injectable hydrogels can be used to deliver drugs in situ over a sustained period of time. We hypothesized that sustained delivery of interleukin-10 (IL-10) following acute kidney injury (AKI) would mitigate the local and systemic proinflammatory cascade induced by AKI and reduce subsequent fibrosis. Wild-type C57BL/6 mice underwent ischemia-reperfusion AKI with avertin anesthesia. Three days later, mice were treated with either hyaluronic acid injectable hydrogel with or without IL-10, or IL-10 suspended in saline, injected under the capsule of the left kidney, or hydrogel with IL-10 injected subcutaneously. Untreated AKI served as controls. Serial in vivo optical imaging tracked the location and degradation of the hydrogel over time. Kidney function was assessed serially. Animals were killed 28 days following AKI and the following were evaluated: serum IL-6, lung inflammation, urine neutrophil gelatinase-associated lipocalin, and renal histology for fibroblast activity, collagen type III deposition and fibrosis via Picrosirius Red staining and second harmonic imaging. Our model shows persistent systemic inflammation, and renal inflammation and fibrosis 28 days following AKI. The hydrogels are biocompatible and reduced serum IL-6 and renal collagen type III 28 days following AKI even when delivered without IL-10. Treatment with IL-10 reduced renal and systemic inflammation, regardless of whether the IL-10 was delivered in a sustained manner via the injectable hydrogel under the left kidney capsule, as a bolus injection via saline under the left kidney capsule, or via the injectable hydrogel subcutaneously. Injectable hydrogels are suitable for local drug delivery following renal injury, are biocompatible, and help mitigate local and systemic inflammation. PMID:26962109

  17. Lateral Transfer of the Denitrification Pathway Genes among Thermus thermophilus Strains▿

    PubMed Central

    Alvarez, Laura; Bricio, Carlos; José Gómez, Manuel; Berenguer, José

    2011-01-01

    Nitrate respiration is a common and strain-specific property in Thermus thermophilus encoded by the nitrate respiration conjugative element (NCE) that can be laterally transferred by conjugation. In contrast, nitrite respiration and further denitrification steps are restricted to a few isolates of this species. These later steps of the denitrification pathway are under the regulatory control of an NCE-encoded transcription factor, but nothing is known about their coding sequences or its putative genetic linkage to the NCE. In this study we examine the genetic linkage between nitrate and nitrite respiration through lateral gene transfer (LGT) assays and describe a cluster of genes encoding the nitrite-nitric oxide respiration in T. thermophilus PRQ25. We show that the whole denitrification pathway can be transferred from the denitrificant strain PRQ25 to an aerobic strain, HB27, and that the genes coding for nitrite and nitric oxide respiration are encoded near the NCE. Sequence data from the draft genome of PRQ25 confirmed these results and allowed us to describe the most compact nor-nir cluster known thus far and to demonstrate the expression and activities of the encoded enzymes in the HB27 denitrificant derivatives obtained by LGT. We conclude that this NCE nor-nir supercluster constitutes a whole denitrification island that can be spread by lateral transfer among Thermus thermophilus strains. PMID:21169443

  18. Evolution and Distribution of the ospC Gene, a Transferable Serotype Determinant of Borrelia burgdorferi

    PubMed Central

    Barbour, Alan G.; Travinsky, Bridgit

    2010-01-01

    Borrelia burgdorferi, an emerging bacterial pathogen, is maintained in nature by transmission from one vertebrate host to another by ticks. One of the few antigens against which mammals develop protective immunity is the highly polymorphic OspC protein, encoded by the ospC gene on the cp26 plasmid. Intragenic recombination among ospC genes is known, but the extent to which recombination extended beyond the ospC locus itself is undefined. We accessed and supplemented collections of DNA sequences of ospC and other loci from ticks in three U.S. regions (the Northeast, the Midwest, and northern California); a total of 839 ospC sequences were analyzed. Three overlapping but distinct populations of B. burgdorferi corresponded to the geographic regions. In addition, we sequenced 99 ospC flanking sequences from different lineages and compared the complete cp26 sequences of 11 strains as well as the cp26 bbb02 loci of 56 samples. Besides recombinations with traces limited to the ospC gene itself, there was evidence of lateral gene transfers that involved (i) part of the ospC gene and one of the two flanks or (ii) the entire ospC gene and different lengths of both flanks. Lateral gene transfers resulted in different linkages between the ospC gene and loci of the chromosome or other plasmids. By acquisition of the complete part or a large part of a novel ospC gene, an otherwise adapted strain would assume a new serotypic identity, thereby being comparatively fitter in an area with a high prevalence of immunity to existing OspC types. PMID:20877579

  19. Inducible long-term gene expression in brain with adeno-associated virus gene transfer.

    PubMed

    Haberman, R P; McCown, T J; Samulski, R J

    1998-12-01

    Recombinant adeno-associated virus (rAAV) vectors hold promise for treating a number of neurological disorders due to the ability to deliver long-term gene expression without toxicity or immune response. Critical to these endeavors will be controlled expression of the therapeutic gene in target cells. We have constructed and tested a dual cassette rAAV vector carrying a reporter gene under the control of the tetracycline-responsive system and the tetracycline transactivator. Transduction in vitro resulted in stable expression from the vector that can be suppressed 20-fold by tetracycline treatment. In vivo experiments, carried out to 6 weeks, demonstrated that vector-transduced expression is sustained until doxycycline administration upon which reporter gene expression is reduced. Moreover, the suppression of vector-driven expression can be reversed by removal of the drug. These studies demonstrate long-term regulated gene expression from rAAV vectors. This system will provide a valuable approach for controlling vector gene expression both in vitro and in vivo. PMID:10023439

  20. Transcriptional Analysis of the Conjugal Transfer Genes of Rickettsia bellii RML 369-C

    PubMed Central

    Heu, Chan C.; Kurtti, Timothy J.; Nelson, Curtis M.; Munderloh, Ulrike G.

    2015-01-01

    Rickettsia bellii is an obligate intracellular bacterium that is one of the few rickettsiae that encode a complete set of conjugative transfer (tra) genes involved in bacterial conjugation and has been shown to exhibit pili-like structures. The reductive genomes of rickettsiae beg the question whether the tra genes are nonfunctional or functioning to enhance the genetic plasticity and biology of rickettsiae. We characterized the transcriptional dynamics of R. bellii tra genes in comparison to genes transcribed stably and above the background level to understand when and at what levels the tra genes are active or whether the tra genes are degenerative. We determined that the best reference genes, out of 10 tested, were methionyl tRNA ligase (metG) or a combination of metG and ribonucleoside diphosphate reductase 2 subunit beta (nrdF), using statistical algorithms from two different programs: Normfinder and BestKeeper. To validate the use of metG with other rickettsial genes exhibiting variable transcriptional patterns we examined its use with sca2 and rickA, genes involved in actin based motility. Both were shown to be up-regulated at different times of replication in Vero cells, showing variable and stable transcription levels of rickA and sca2, respectively. traATi was up-regulated at 72 hours post inoculation in the tick cell line ISE6, but showed no apparent changes in the monkey cell line Vero and mouse cell line L929. The transcription of tra genes was positively correlated with one another and up-regulated from 12 to 72 hours post inoculation (HPI) when compared to RBE_0422 (an inactivated transposase-derivative found within the tra cluster). Thus, the up-regulation of the tra genes indicated that the integrity and activity of each gene were intact and may facilitate the search for the optimal conditions necessary to demonstrate conjugation in rickettsiae. PMID:26352829

  1. Horizontal gene transfers and cell fusions in microbiology, immunology and oncology (Review).

    PubMed

    Sinkovics, Joseph G

    2009-09-01

    Evolving young genomes of archaea, prokaryota and unicellular eukaryota were wide open for the acceptance of alien genomic sequences, which they often preserved and vertically transferred to their descendants throughout three billion years of evolution. Established complex large genomes, although seeded with ancestral retroelements, have come to regulate strictly their integrity. However, intruding retroelements, especially the descendents of Ty3/Gypsy, the chromoviruses, continue to find their ways into even the most established genomes. The simian and hominoid-Homo genomes preserved and accommodated a large number of endogenous retroviral genomic segments. These retroelements may mature into exogenous retroviruses, or into functional new genes. Phages and viruses have been instrumental in incorporating and transferring host cell genes. These events profoundly influenced and altered the course of evolution. Horizontal (lateral) gene transfers (HGT) overwhelmed the genomes of the ancient protocells and the evolving unicellular microorganisms, actually leading to their Cambrian explosion. While the rigidly organized genomes of multicellular organisms increasingly resist H/LGT, de-differentiated cells assuming the metabolism of their onto- or phylogenetic ancestors, open up widely to the practice of H/LGT by direct transfer, or to transfers mediated by viruses, or by cell fusions. This activity is intensified in malignantly transformed cells, thus rendering these subjects receptive to therapy with oncolytic viruses and with viral vectors of tumor-suppressive or immunogenic genetic materials. Naturally formed hybrids of dendritic and tumor cells are often tolerogenic, whereas laboratory products of these unisons may be immunogenic in the hosts of origin. As human breast cancer stem cells are induced by a treacherous class of CD8+ T cells to undergo epithelial to mesenchymal (ETM) transition and to yield to malignant transformation by the omnipresent proto

  2. Plant–Agrobacterium interaction mediated by ethylene and super-Agrobacterium conferring efficient gene transfer

    PubMed Central

    Nonaka, Satoko; Ezura, Hiroshi

    2014-01-01

    Agrobacterium tumefaciens has a unique ability to transfer genes into plant genomes. This ability has been utilized for plant genetic engineering. However, the efficiency is not sufficient for all plant species. Several studies have shown that ethylene decreased the Agrobacterium-mediated transformation frequency. Thus, A. tumefaciens with an ability to suppress ethylene evolution would increase the efficiency of Agrobacterium-mediated transformation. Some studies showed that plant growth-promoting rhizobacteria (PGPR) can reduce ethylene levels in plants through 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase, which cleaves the ethylene precursor ACC into α-ketobutyrate and ammonia, resulting in reduced ethylene production. The whole genome sequence data showed that A. tumefaciens does not possess an ACC deaminase gene in its genome. Therefore, providing ACC deaminase activity to the bacteria would improve gene transfer. As expected, A. tumefaciens with ACC deaminase activity, designated as super-Agrobacterium, could suppress ethylene evolution and increase the gene transfer efficiency in several plant species. In this review, we summarize plant–Agrobacterium interactions and their applications for improving Agrobacterium-mediated genetic engineering techniques via super-Agrobacterium. PMID:25520733

  3. Cell-specific expression of the carrot EP2 lipid transfer protein gene.

    PubMed Central

    Sterk, P; Booij, H; Schellekens, G A; Van Kammen, A; De Vries, S C

    1991-01-01

    A cDNA corresponding to a 10-kD protein, designated extracellular protein 2 (EP2), that is secreted by embryogenic cell cultures of carrot was obtained by expression screening. The derived protein sequence and antisera against heterologous plant lipid transfer proteins identified the EP2 protein as a lipid transfer protein. Protein gel blot analysis showed that the EP2 protein is present in cell walls and conditioned medium of cell cultures. RNA gel blot analysis revealed that the EP2 gene is expressed in embryogenic cell cultures, the shoot apex of seedlings, developing flowers, and maturing seeds. In situ hybridization showed expression of the EP2 gene in protoderm cells of somatic and zygotic embryos and transient expression in epidermis cells of leaf primordia and all flower organs. In the shoot apical meristem, expression is found in the tunica and lateral zone. In maturing seeds, the EP2 gene is expressed in the outer epidermis of the integument, the seed coat, and the pericarp epidermis, as well as transiently in between both mericarps. Based on the extracellular location of the EP2 protein and the expression pattern of the encoding gene, we propose a role for plant lipid transfer proteins in the transport of cutin monomers through the extracellular matrix to sites of cutin synthesis. PMID:1822991

  4. Pigment-cell-specific genes from fibroblasts are transactivated after chromosomal transfer into melanoma cells

    SciTech Connect

    Powers, T.P.; Davidson, R.L.; Shows, T.B.

    1994-02-01

    Human and mouse fibroblast chromosomes carrying tyrosinase or b-locus genes were introduced, by microcell hybridization, into pigmented Syrian hamster melanoma cells, and the microcell hybrids were tested for transactivation of the fibroblast tyrosinase and b-locus genes. By using species-specific PCR amplification to distinguish fibroblast and melanoma cDNAs, it was demonstrated that the previously silent fibroblast tyrosinase and b-locus genes were transactivated following chromosomal transfer into pigmented melanoma cells. However, transactivation of the mouse fibroblast tyrosinase gene was unstable in microcell hybrid subclones and possibly dependent on a second fibroblast locus that could have segregated in the subclones. This second locus was not necessary for transactivation of the fibroblast b-locus gene, thus demonstrating noncoordinate transactivation of fibroblast tyrosinase and b-locus genes. Transactivation of the fibroblast tyrosinase gene in microcell hybrids apparently is dependent on the absence of a putative fibroblast extinguisher locus for tyrosinase gene expression, which presumably is responsible for the extinction of pigmentation in hybrids between karyotypically complete fibroblasts and melanoma cells. 46 refs., 5 figs., 2 tabs.

  5. Enterococcus faecalis Gene Transfer under Natural Conditions in Municipal Sewage Water Treatment Plants†

    PubMed Central

    Marcinek, Herbert; Wirth, Reinhard; Muscholl-Silberhorn, Albrecht; Gauer, Matthias

    1998-01-01

    The ability of Enterococcus faecalis to transfer various genetic elements under natural conditions was tested in two municipal sewage water treatment plants. Experiments in activated sludge basins of the plants were performed in a microcosm which allowed us to work under sterile conditions; experiments in anoxic sludge digestors were performed in dialysis bags. We used the following naturally occurring genetic elements: pAD1 and pIP1017 (two so-called sex pheromone plasmids with restricted host ranges, which are transferred at high rates under laboratory conditions); pIP501 (a resistance plasmid possessing a broad host range for gram-positive bacteria, which is transferred at low rates under laboratory conditions); and Tn916 (a conjugative transposon which is transferred under laboratory conditions at low rates to gram-positive bacteria and at very low rates to gram-negative bacteria). The transfer rate between different strains of E. faecalis under natural conditions was, compared to that under laboratory conditions, at least 105-fold lower for the sex pheromone plasmids, at least 100-fold lower for pIP501, and at least 10-fold lower for Tn916. In no case was transfer from E. faecalis to another bacterial species detected. By determining the dependence of transfer rates for pIP1017 on bacterial concentration and extrapolating to actual concentrations in the sewage water treatment plant, we calculated that the maximum number of transfer events for the sex pheromone plasmids between different strains of E. faecalis in the municipal sewage water treatment plant of the city of Regensburg ranged from 105 to 108 events per 4 h, indicating that gene transfer should take place under natural conditions. PMID:9464401

  6. Phylogeographic reconstruction of a bacterial species with high levels of lateral gene transfer

    USGS Publications Warehouse

    Pearson, T.; Giffard, P.; Beckstrom-Sternberg, S.; Auerbach, R.; Hornstra, H.; Tuanyok, A.; Price, E.P.; Glass, M.B.; Leadem, B.; Beckstrom-Sternberg, J. S.; Allan, G.J.; Foster, J.T.; Wagner, D.M.; Okinaka, R.T.; Sim, S.H.; Pearson, O.; Wu, Z.; Chang, J.; Kaul, R.; Hoffmaster, A.R.; Brettin, T.S.; Robison, R.A.; Mayo, M.; Gee, J.E.; Tan, P.; Currie, B.J.; Keim, P.

    2009-01-01

    Background: Phylogeographic reconstruction of some bacterial populations is hindered by low diversity coupled with high levels of lateral gene transfer. A comparison of recombination levels and diversity at seven housekeeping genes for eleven bacterial species, most of which are commonly cited as having high levels of lateral gene transfer shows that the relative contributions of homologous recombination versus mutation for Burkholderia pseudomallei is over two times higher than for Streptococcus pneumoniae and is thus the highest value yet reported in bacteria. Despite the potential for homologous recombination to increase diversity, B. pseudomallei exhibits a relative lack of diversity at these loci. In these situations, whole genome genotyping of orthologous shared single nucleotide polymorphism loci, discovered using next generation sequencing technologies, can provide very large data sets capable of estimating core phylogenetic relationships. We compared and searched 43 whole genome sequences of B. pseudomallei and its closest relatives for single nucleotide polymorphisms in orthologous shared regions to use in phylogenetic reconstruction. Results: Bayesian phylogenetic analyses of >14,000 single nucleotide polymorphisms yielded completely resolved trees for these 43 strains with high levels of statistical support. These results enable a better understanding of a separate analysis of population differentiation among >1,700 B. pseudomallei isolates as defined by sequence data from seven housekeeping genes. We analyzed this larger data set for population structure and allele sharing that can be attributed to lateral gene transfer. Our results suggest that despite an almost panmictic population, we can detect two distinct populations of B. pseudomallei that conform to biogeographic patterns found in many plant and animal species. That is, separation along Wallace's Line, a biogeographic boundary between Southeast Asia and Australia. Conclusion: We describe an

  7. Optimizing in vivo gene transfer into mouse corpus cavernosum by use of surface electroporation

    PubMed Central

    Song, Kang-Moon; Choi, Min Ji; Kwon, Mi-Hye; Ghatak, Kalyan; Park, Soo-Hwan; Ryu, Dong-Soo; Ryu, Ji-Kan

    2015-01-01

    Purpose Electroporation is known to enhance the efficiency of gene transfer through a transient increase in cell membrane permeability. The aim of this study was to determine the optimal conditions for in vivo electroporation-mediated gene delivery into mouse corpus cavernosum. Materials and Methods Diabetes was induced in C57BL/6 mice by intraperitoneal injections of streptozotocin. After intracavernous injection of pCMV-Luc (100 µg/40 µL), different electroporation settings (5-50 V, 8-16 pulses with a duration of 40-100 ms) were applied to the penis to establish the optimal conditions for electroporation. Gene expression was evaluated by luciferase assay. We also assessed the undesired consequences of electroporation by visual inspection and hematoxylin-eosin staining of penile tissue. Results Electroporation profoundly induced gene expression in the corpus cavernosum tissue of normal mice in a voltage-dependent manner. We observed electrical burn scars in the penis of normal mice who received electroporation with eight 40-ms pulses at a voltage of 50 V and sixteen 40-ms pulses, eight 100-ms pulses, and sixteen 100-ms pulses at a voltage of 30 V. No detectable burn scars were noted in normal mice stimulated with eight 40-ms pulses at a voltage of 30 V. Electroporation also significantly induced gene expression in diabetic mice stimulated with 40-ms pulse at a voltage of 30 V without injury to the penis. Conclusions We have established the optimal electroporation conditions for maximizing gene transfer into the corpus cavernosum of mice while avoiding damage to the erectile tissue. The electroporation-mediated gene delivery technique will be a valuable tool for gene therapy in the field of erectile dysfunction. PMID:25763123

  8. Bacterial α2-macroglobulins: colonization factors acquired by horizontal gene transfer from the metazoan genome?

    PubMed Central

    Budd, Aidan; Blandin, Stephanie; Levashina, Elena A; Gibson, Toby J

    2004-01-01

    Background Invasive bacteria are known to have captured and adapted eukaryotic host genes. They also readily acquire colonizing genes from other bacteria by horizontal gene transfer. Closely related species such as Helicobacter pylori and Helicobacter hepaticus, which exploit different host tissues, share almost none of their colonization genes. The protease inhibitor α2-macroglobulin provides a major metazoan defense against invasive bacteria, trapping attacking proteases required by parasites for successful invasion. Results Database searches with metazoan α2-macroglobulin sequences revealed homologous sequences in bacterial proteomes. The bacterial α2-macroglobulin phylogenetic distribution is patchy and violates the vertical descent model. Bacterial α2-macroglobulin genes are found in diverse clades, including purple bacteria (proteobacteria), fusobacteria, spirochetes, bacteroidetes, deinococcids, cyanobacteria, planctomycetes and thermotogae. Most bacterial species with bacterial α2-macroglobulin genes exploit higher eukaryotes (multicellular plants and animals) as hosts. Both pathogenically invasive and saprophytically colonizing species possess bacterial α2-macroglobulins, indicating that bacterial α2-macroglobulin is a colonization rather than a virulence factor. Conclusions Metazoan α2-macroglobulins inhibit proteases of pathogens. The bacterial homologs may function in reverse to block host antimicrobial defenses. α2-macroglobulin was probably acquired one or more times from metazoan hosts and has then spread widely through other colonizing bacterial species by more than 10 independent horizontal gene transfers. yfhM-like bacterial α2-macroglobulin genes are often found tightly linked with pbpC, encoding an atypical peptidoglycan transglycosylase, PBP1C, that does not function in vegetative peptidoglycan synthesis. We suggest that YfhM and PBP1C are coupled together as a periplasmic defense and repair system. Bacterial α2-macroglobulins might

  9. Gene Transfer by Guanidinium-Cholesterol Cationic Lipids into Airway Epithelial Cells in vitro and in vivo

    NASA Astrophysics Data System (ADS)

    Oudrhiri, Noufissa; Vigneron, Jean-Pierre; Peuchmaur, Michel; Leclerc, Tony; Lehn, Jean-Marie; Lehn, Pierre

    1997-03-01

    Synthetic vectors represent an attractive alternative approach to viral vectors for gene transfer, in particular into airway epithelial cells for lung-directed gene therapy for cystic fibrosis. Having recently found that guanidinium-cholesterol cationic lipids a