<em>L. donovani</em> infections (visceral and post kala-azar dermal leishmaniases) are characterized by infection-induced reversible immune suppression. Autoimmunity is a well-documented phenomenon among p...<em>L. donovani</em> infections (visceral and post kala-azar dermal leishmaniases) are characterized by infection-induced reversible immune suppression. Autoimmunity is a well-documented phenomenon among patients with primary immune deficiencies. This study aimed to study auto-immune phenomena accompanying <em>L. donovani</em> infections. In a prospective case-controlled study and following informed consent, 155 individuals with visceral leishmaniasis (VL;<em>n</em> = 62), post kala-azar dermal leishmaniasis (PKDL;<em>n</em> = 31) and apparently healthy volunteers (<em>n</em> = 62) were recruited. Sera antinuclear (ANA), anti-dsDNA, anti-thyroid peroxidase (TPO), anti-smooth muscles (ASMA) and F-actin auto-antibodies were measured using ELISA and indirect immune-fluorescence assay. The mean ages of VL, PDKL patients and apparently healthy volunteers were: 17.5 ± 12.5, 15.0 ± 7.0 and 17.5 ± 9.5 years with Male:Female ratios of 2:0, 1:2 and 1:5 respectively. Significantly high frequencies of F-actin (74.2%;46/62) and ASMA (50%;31/62) auto-antibodies were seen among VL patients (<strong><em>p</em> = 0.003</strong>, <strong><em>p</em> = 0.001</strong>) compared to apparently healthy volunteers. Likewise, significantly high frequencies of F-actin (64.5%;20/31;<strong><em>p</em> = 0.001</strong>), ASMA (42%;13/31;<strong><em>p</em> = 0.003</strong>), ANA (36%;11/31;<strong><em>p</em> = 0.00</strong><strong>1</strong>) and anti-dsDNA (16%;5/31;<strong><em>p</em> = 0.01</strong>) auto-antibodies were seen among PKDL patients. Development of tissue-based autoantibodies in <em>L. donovani</em> infections probably indicates loss of peripheral tolerance with activation of circulating auto-reactive T and B cells probably contributing to disease pathogenesis (increased bilirubin/liver enzymes, prolonged QT interval/arrythmias and blood cytopenias). In conclusion, <em>L. donovani</em> infection-induced immune suppression with development of tissue-based auto-antibodies is prevalent among Sudanese patients with VL and PKDL leishmaniases and contributes to some aspects of the disease pathogenesis.展开更多
<span style="font-family:Verdana;">L-Theanine (</span><i><span style="font-family:Verdana;">γ</span></i><span style="font-family:Verdana;">-glut...<span style="font-family:Verdana;">L-Theanine (</span><i><span style="font-family:Verdana;">γ</span></i><span style="font-family:Verdana;">-glutamylethylamide) is a naturally occurring amino acid derivative known to have several beneficial physiological effects as a diet supplement, and to give an umami taste when used as a food additive. The compound is industrially produced by </span><i><span style="font-family:Verdana;">γ</span></i><span style="font-family:Verdana;">-glutamyltranspeptidase from </span><i><span style="font-family:Verdana;">Pseudomonas nitroreducens</span></i><span style="font-family:Verdana;"> (PnGGT). Using recombinant PnGGT, we have shown previously that Trp385, Phe417, and Trp525 are key amino acid residues for recognition of acceptor substrates at the PnGGT active site. Here, we demonstrate that a recombinant W525D mutant of PnGGT produces L-theanine from ethylamine and L-glutamine more efficiently than wild-type PnGGT, attributable to an increased ratio of transfer activity to hydrolysis activity. An efficient production of L-theanine was achieved by immobilizing </span><i><span style="font-family:Verdana;">Escherichia coli</span></i><span style="font-family:Verdana;"> cells expressing the W525D PnGGT mutant (</span><i><span style="font-family:Verdana;">E. coli</span></i><span style="font-family:Verdana;">-W525D) using 2% alginate as the supporting material. The highest L-theanine production using immobilized </span><i><span style="font-family:Verdana;">E. coli</span></i><span style="font-family:Verdana;">-W525D, representing a conversion rate of 90%, w</span><span style="font-family:Verdana;">as</span><span style="font-family:Verdana;"> achieved in optimal reaction conditions of pH 10, 40<span style="white-space:nowrap;">°</span></span><span style="font-family:;" "=""><span><span style="font-family:Verdana;">C, and a substrate molar ratio of L-glutamine to ethylamine of 1:10. The immobilized </span><i><span style="font-family:Verdana;">E. coli</span></i><span style="font-family:Verdana;">-W525D retains 85% and 78% relative activity after storage for a month at 4<span style="white-space:nowrap;">°</span></span></span><span><span style="font-family:Verdana;">C and room temperature, respectively. Immobilized </span><i><span style="font-family:Verdana;">E. coli</span></i><span style="font-family:Verdana;">-W525D thus has strong potential for use in the future commercial production of L-theanine on a large scale.</span></span></span>展开更多
将1.4kb Class I patatin基因的5′侧翼区与GUS基因融合,构建了双元表达载体pPATIs(含patatin部分信号顺序)和pPATI(不含patatin部分信号顺序)。pPATI通过基因枪介导在块茎切片中获得了瞬间表达。以上建构物通过农杆菌介导转入了马铃薯...将1.4kb Class I patatin基因的5′侧翼区与GUS基因融合,构建了双元表达载体pPATIs(含patatin部分信号顺序)和pPATI(不含patatin部分信号顺序)。pPATI通过基因枪介导在块茎切片中获得了瞬间表达。以上建构物通过农杆菌介导转入了马铃薯品种Desiree。X-Gluc染色(PATIs不能染色)及PCR结果证实已获得转基因植株。利用离体块茎诱导系统,GUS的表达进一步用荧光进行定量检测,结果显示,PATI-GUS的转基因植株中GUS比活性均以块茎明显高于茎段,达10-20倍。蔗糖浓度的升高,PATI-GUS植株中的GUS比活性无明显变化,与前人报道有不同。此外,光照促进PATI-GUS的表达。展开更多
文摘<em>L. donovani</em> infections (visceral and post kala-azar dermal leishmaniases) are characterized by infection-induced reversible immune suppression. Autoimmunity is a well-documented phenomenon among patients with primary immune deficiencies. This study aimed to study auto-immune phenomena accompanying <em>L. donovani</em> infections. In a prospective case-controlled study and following informed consent, 155 individuals with visceral leishmaniasis (VL;<em>n</em> = 62), post kala-azar dermal leishmaniasis (PKDL;<em>n</em> = 31) and apparently healthy volunteers (<em>n</em> = 62) were recruited. Sera antinuclear (ANA), anti-dsDNA, anti-thyroid peroxidase (TPO), anti-smooth muscles (ASMA) and F-actin auto-antibodies were measured using ELISA and indirect immune-fluorescence assay. The mean ages of VL, PDKL patients and apparently healthy volunteers were: 17.5 ± 12.5, 15.0 ± 7.0 and 17.5 ± 9.5 years with Male:Female ratios of 2:0, 1:2 and 1:5 respectively. Significantly high frequencies of F-actin (74.2%;46/62) and ASMA (50%;31/62) auto-antibodies were seen among VL patients (<strong><em>p</em> = 0.003</strong>, <strong><em>p</em> = 0.001</strong>) compared to apparently healthy volunteers. Likewise, significantly high frequencies of F-actin (64.5%;20/31;<strong><em>p</em> = 0.001</strong>), ASMA (42%;13/31;<strong><em>p</em> = 0.003</strong>), ANA (36%;11/31;<strong><em>p</em> = 0.00</strong><strong>1</strong>) and anti-dsDNA (16%;5/31;<strong><em>p</em> = 0.01</strong>) auto-antibodies were seen among PKDL patients. Development of tissue-based autoantibodies in <em>L. donovani</em> infections probably indicates loss of peripheral tolerance with activation of circulating auto-reactive T and B cells probably contributing to disease pathogenesis (increased bilirubin/liver enzymes, prolonged QT interval/arrythmias and blood cytopenias). In conclusion, <em>L. donovani</em> infection-induced immune suppression with development of tissue-based auto-antibodies is prevalent among Sudanese patients with VL and PKDL leishmaniases and contributes to some aspects of the disease pathogenesis.
文摘<span style="font-family:Verdana;">L-Theanine (</span><i><span style="font-family:Verdana;">γ</span></i><span style="font-family:Verdana;">-glutamylethylamide) is a naturally occurring amino acid derivative known to have several beneficial physiological effects as a diet supplement, and to give an umami taste when used as a food additive. The compound is industrially produced by </span><i><span style="font-family:Verdana;">γ</span></i><span style="font-family:Verdana;">-glutamyltranspeptidase from </span><i><span style="font-family:Verdana;">Pseudomonas nitroreducens</span></i><span style="font-family:Verdana;"> (PnGGT). Using recombinant PnGGT, we have shown previously that Trp385, Phe417, and Trp525 are key amino acid residues for recognition of acceptor substrates at the PnGGT active site. Here, we demonstrate that a recombinant W525D mutant of PnGGT produces L-theanine from ethylamine and L-glutamine more efficiently than wild-type PnGGT, attributable to an increased ratio of transfer activity to hydrolysis activity. An efficient production of L-theanine was achieved by immobilizing </span><i><span style="font-family:Verdana;">Escherichia coli</span></i><span style="font-family:Verdana;"> cells expressing the W525D PnGGT mutant (</span><i><span style="font-family:Verdana;">E. coli</span></i><span style="font-family:Verdana;">-W525D) using 2% alginate as the supporting material. The highest L-theanine production using immobilized </span><i><span style="font-family:Verdana;">E. coli</span></i><span style="font-family:Verdana;">-W525D, representing a conversion rate of 90%, w</span><span style="font-family:Verdana;">as</span><span style="font-family:Verdana;"> achieved in optimal reaction conditions of pH 10, 40<span style="white-space:nowrap;">°</span></span><span style="font-family:;" "=""><span><span style="font-family:Verdana;">C, and a substrate molar ratio of L-glutamine to ethylamine of 1:10. The immobilized </span><i><span style="font-family:Verdana;">E. coli</span></i><span style="font-family:Verdana;">-W525D retains 85% and 78% relative activity after storage for a month at 4<span style="white-space:nowrap;">°</span></span></span><span><span style="font-family:Verdana;">C and room temperature, respectively. Immobilized </span><i><span style="font-family:Verdana;">E. coli</span></i><span style="font-family:Verdana;">-W525D thus has strong potential for use in the future commercial production of L-theanine on a large scale.</span></span></span>
文摘将1.4kb Class I patatin基因的5′侧翼区与GUS基因融合,构建了双元表达载体pPATIs(含patatin部分信号顺序)和pPATI(不含patatin部分信号顺序)。pPATI通过基因枪介导在块茎切片中获得了瞬间表达。以上建构物通过农杆菌介导转入了马铃薯品种Desiree。X-Gluc染色(PATIs不能染色)及PCR结果证实已获得转基因植株。利用离体块茎诱导系统,GUS的表达进一步用荧光进行定量检测,结果显示,PATI-GUS的转基因植株中GUS比活性均以块茎明显高于茎段,达10-20倍。蔗糖浓度的升高,PATI-GUS植株中的GUS比活性无明显变化,与前人报道有不同。此外,光照促进PATI-GUS的表达。
