Biomedical subjects

Michael Schiff

Publications and source records attributed to Michael Schiff.

27 records · Page 2Linked to original sources

PubMed2003

Two MAPK cascades, NPR1, and TGA transcription factors play a role in Pto-mediated disease resistance in tomato.

The tomato Pto kinase confers resistance to the causative agent of bacterial speck disease, Pseudomonas syringae pv. tomato, by recognizing the pathogen effector proteins AvrPto or AvrPtoB. Pto-mediated resistance requires multiple signal transduction pathways and has been shown to activate many defense responses including an oxidative burst, rapid changes in the expression of over 400 genes, and localized cell death. We have tested the role in Pto-mediated resistance in tomato of a set of 21 genes from other species known to be involved in defense-related signaling. Expression of each gene was suppressed by virus-induced gene silencing (VIGS) and the effect on disease symptoms and bacterial growth during the tomato-Pseudomonas incompatible interaction was determined. We found that Pto-mediated resistance was compromised by silencing of genes encoding two mitogen-activated protein (MAP) kinase kinases, MEK1 and MEK2, two MAP kinases, NTF6 and wound-induced protein kinase (WIPK), a key regulator of systemic acquired resistance (SAR), NPR1, and two transcription factors, TGA1a and TGA2.2. A lesser impact on Pto-mediated resistance was observed in plants silenced for RAR1 and COI1. The identification of nine genes that play a role in resistance to bacterial speck disease both advances our knowledge of Pto signal transduction and demonstrates the conservation of many defense signaling components among diverse plant species.

Base Sequence↗

PubMed2003

Virus-induced gene silencing.

In the postgenomic era, large-scale functional genomic approaches are necessary for converting sequence information into functional information. A para-genetic approach, called virus-induced gene silencing (VIGS), offers a rapid means of gaining insight into gene function in plants. VIGS system could be used to suppress endogenous gene expression by infecting plants with a recombinant virus vector (VIGS vector) carrying host-derived sequence. Here, we describe the use of tobacco rattle virus (TRV)-based VIGS technique to study gene function in Nicotiana benthamiana and tomato.

Gene Silencing↗

PubMed2002

Treatment of rheumatoid arthritis with anakinra, a recombinant human interleukin-1 receptor antagonist, in combination with methotrexate: results of a twenty-four-week, multicenter, randomized, double-blind, placebo-controlled trial.

OBJECTIVE: To evaluate the efficacy and safety of anakinra in combination with methotrexate (MTX) in patients with active rheumatoid arthritis (RA). METHODS: Patients with moderate-to-severe active RA who were receiving MTX for 6 consecutive months, with stable doses for > or = 3 months (those with disease duration of >6 months but <12 years) were randomized into 6 groups: placebo or 0.04, 0.1, 0.4, 1.0, or 2.0 mg/kg of anakinra administered in a single, daily, subcutaneous injection. The primary efficacy end point was the proportion of subjects who met the American College of Rheumatology 20% improvement criteria (attained an ACR20 response) at week 12. RESULTS: A total of 419 patients were randomized in the study. Patient demographics and disease status were similar in the 6 treatment groups. The ACR20 responses at week 12 in the 5 active treatment plus MTX groups demonstrated a statistically significant (P = 0.001) dose-response relationship compared with the ACR20 response in the placebo plus MTX group. The ACR20 response rate in the anakinra 1.0-mg/kg (46%; P = 0.001) and 2.0-mg/kg (38%; P = 0.007) dose groups was significantly greater than that in the placebo group (19%). The ACR20 responses at 24 weeks were consistent with those at 12 weeks. Similar improvements in anakinra-treated subjects were noted in individual ACR components, erythrocyte sedimentation rate, onset of ACR20 response, sustainability of ACR20 response, and magnitude of ACR response. Anakinra was safe and well tolerated. Injection site reaction was the most frequently noted adverse event, and this led to premature study withdrawal in 7% (1.0-mg/kg group) to 10% (2.0-mg/kg group) of patients receiving higher doses. CONCLUSION: In patients with persistently active RA, the combination of anakinra and MTX was safe and well tolerated and provided significantly greater clinical benefit than MTX alone.

Antibodies↗

PubMed2002

Effects of prasterone on corticosteroid requirements of women with systemic lupus erythematosus: a double-blind, randomized, placebo-controlled trial.

OBJECTIVE: To evaluate whether treatment with prasterone (dehydroepiandrosterone [DHEA]) would allow the dosage of prednisone (or an equivalent corticosteroid) to be reduced to < or = 7.5 mg/day for 2 months or longer while maintaining stable or reduced disease activity in steroid-dependent women with systemic lupus erythematosus (SLE). METHODS: In a double-blind, randomized trial, 191 female SLE patients receiving prednisone (10-30 mg/day) were treated daily with either placebo, 100 mg of oral prasterone (an adrenal androgen), or 200 mg of oral prasterone for 7-9-months. At monthly intervals, corticosteroid dosages were reduced by algorithm in patients whose SLE Disease Activity Index (SLEDAI) score was stable or improved. Patients for whom a sustained reduction in the dosage of prednisone (< or = 7.5 mg/day) was achieved for at least the last 2 months of the 7-9-month treatment period were classified as responders. RESULTS: Response rates were 41% in the placebo group, 44% in the 100-mg prasterone group, and 55% in the 200-mg group (P = 0.110, 200 mg versus placebo). Among the 137 subjects (45 in the placebo group, 47 in the 100-mg group, and 45 in the 200-mg group) who had active disease at baseline (defined as SLEDAI score >2), 29%, 38%, and 51%, respectively, were responders (P = 0.031 for 200 mg prasterone versus placebo). Acne was the most common adverse event but was generally mild. Clinical and laboratory changes primarily reflected androgenic effects of prasterone. CONCLUSION: Among women with lupus disease activity, reducing the dosage of prednisone to < or = 7.5 mg/day for a sustained period of time while maintaining stabilization or a reduction of disease activity was possible in a significantly greater proportion of patients treated with oral prasterone, 200 mg once daily, compared with patients treated with placebo.

Administration, Oral↗

PubMed2002

Efficacy of tacrolimus in rheumatoid arthritis patients who have been treated unsuccessfully with methotrexate: a six-month, double-blind, randomized, dose-ranging study.

OBJECTIVE: To assess the efficacy, safety, and optimal dose of tacrolimus monotherapy in patients with rheumatoid arthritis (RA). METHODS: This phase II, randomized, double-blind, placebo-controlled monotherapy study was set in 12 community sites and 9 university-based sites. Two hundred sixty-eight patients with RA who were resistant to or intolerant of methotrexate (mean dose 15.2 mg/week) and had active disease for at least 6 months (mean tender joint count 28.2, mean erythrocyte sedimentation rate 46.5 mm/hour) were randomized to receive treatment after discontinuation of methotrexate. Those who received at least 1 dose of tacrolimus were analyzed; 141 completed the study. Stable dosages of nonsteroidal antiinflammatory drugs and low-dose prednisone were allowed during treatment. All patients were given 1, 3, or 5 mg of tacrolimus or placebo once daily for 24 weeks. The American College of Rheumatology definition of 20% improvement (ACR20) and the tender and swollen joint counts at the end of treatment were the primary outcomes. RESULTS: ACR20 response rates demonstrated a clear dose response. The ACR20 response was observed in 15.5% of patients receiving placebo (95% confidence interval [95% CI] 7.1-23.9%), 29% of the 1 mg tacrolimus group (95% CI 18.3-39.7%) (P < 0.058); 34.4% of the 3 mg group (95% CI 22.7-46.0%) (P < 0.013), and 50% of the 5 mg group (95% CI 37.8-62.3%) (P < or = 0.001). The tender joint count improved statistically significantly in all tacrolimus groups. The swollen joint count, physical function, and patient-assessed pain improved statistically significantly in the 3 mg and 5 mg groups. The incidence of creatinine elevation > or =40% above baseline levels increased in a dose-dependent manner. Dropout rates were high (41-59%) and were more common for inefficacy in the placebo patients (71.4%), whereas they were more common for toxicity in the high-dose tacrolimus groups (31-33%). Discontinuation for creatinine elevation occurred in the 3 mg (3.1%) and 5 mg (10.9%) tacrolimus groups. CONCLUSION: Tacrolimus improved disease activity in methotrexate-resistant or -intolerant patients with RA. A dose response was observed when efficacy and toxicity were assessed at different doses. The optimal dose of tacrolimus appears to be >1 mg but < or=3 mg daily.

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed2002

Tobacco Rar1, EDS1 and NPR1/NIM1 like genes are required for N-mediated resistance to tobacco mosaic virus.

The tobacco N gene confers resistance to tobacco mosaic virus (TMV) and encodes a Toll-interleukin-1 receptor/nucleotide binding site/leucine-rich repeat (TIR-NBS-LRR) class protein. We have developed and used a tobacco rattle virus (TRV) based virus induced gene silencing (VIGS) system to investigate the role of tobacco candidate genes in the N-mediated signalling pathway. To accomplish this we generated transgenic Nicotiana benthamiana containing the tobacco N gene. The transgenic lines exhibit hypersensitive response (HR) to TMV and restrict virus spread to the inoculated site. This demonstrates that the tobacco N gene can confer resistance to TMV in heterologous N. benthamiana. We have used this line to study the role of tobacco Rar1-, EDS1-, and NPR1/NIM1- like genes in N-mediated resistance to TMV using a TRV based VIGS approach. Our VIGS analysis suggests that these genes are required for N function. EDS1-like gene requirement for the N function suggests that EDS1 could be a common component of bacterial, fungal and viral resistance signalling mediated by the TIR-NBS-LRR class of resistance proteins. Requirement of Rar1- like gene for N-mediated resistance to TMV and some powdery mildew resistance genes in barley provide the first example of converging points in the disease resistance signalling pathways mediated by TIR-NBS-LRR and CC-NBS-LRR proteins. The TRV based VIGS approach as described here to study N-mediated resistance signalling will be useful for the analysis of not only disease resistance signalling pathways but also of other signalling pathways in genetically intractable plant systems.

Amino Acid Sequence↗

PubMed2002

Virus-induced gene silencing in tomato.

We have previously demonstrated that a tobacco rattle virus (TRV)-based vector can be used in virus-induced gene silencing (VIGS) to study gene function in Nicotiana benthamiana. Here we show that recombinant TRV infects tomato plants and induces efficient gene silencing. Using this system, we suppressed the PDS, CTR1 and CTR2 genes in tomato. Suppression of CTR1 led to a constitutive ethylene response phenotype and up-regulation of an ethylene response gene, CHITINASE B. This phenotype is similar to Arabidopsis ctr1 mutant plants. We have constructed a modified TRV vector based on the GATEWAY recombination system, allowing restriction- and ligation-free cloning. Our results show that tomato expressed sequence tags (ESTs) can easily be cloned into this modified vector using a single set of primers. Using this vector, we have silenced RbcS and an endogenous gene homologous to the tomato EST cLED3L14. In the future, this modified vector system will facilitate large-scale functional analysis of tomato ESTs.

Carrier Proteins↗

PubMed2002

Role of SCF ubiquitin-ligase and the COP9 signalosome in the N gene-mediated resistance response to Tobacco mosaic virus.

The tobacco N gene confers resistance to Tobacco mosaic virus (TMV) and encodes a toll-interleukin-1 receptor/nucleotide binding/Leu-rich repeat class protein. Recent evidence indicates that the Nicotiana benthamiana Rar1 gene (NbRar1), which encodes a protein with a zinc finger motif called CHORD (Cys- and His-rich domain), is required for the function of N. To investigate the role of NbRar1 in plant defense, we identified its interaction partners. We show that the NbRar1 protein interacts with NbSGT1, a highly conserved component of the SCF (Skp1/Cullin/F-box protein)-type E3 ubiquitin ligase complex involved in protein degradation. In addition, we show that NbSGT1 interacts with NbSKP1. Suppression of NbSGT1 and NbSKP1 shows that these genes play an important role in the N-mediated resistance response to TMV. Both NbRar1 and NbSGT1 associate with the COP9 signalosome, another multiprotein complex involved in protein degradation via the ubiquitin-proteasome pathway. Silencing of the NbCOP9 signalosome also compromises N-mediated resistance to TMV. Our results reveal new roles for SCF and the COP9 signalosome in plant defense signaling.

Amino Acid Motifs↗

PubMedDate not supplied

Aberrant crypt foci.

Colon cancer evolves through epithelial cell deregulation and inappropriate proliferation. These histopathological characteristics are exemplified in the biochemical, immunohistochemical, genetic and epigenetic elements detected within colonic mucosa. Early detection is paramount for the prevention of colon cancer deaths. Aberrant crypt foci (ACF) are thought to be the earliest identifiable neoplastic lesions in the colon carcinogenetic model. The progression of ACF to polyp and, subsequently, to cancer parallels the accumulation of several biochemical alterations and mutations whereby a small fraction of ACF evolve to colon cancer. Recent data indicate that, not uncommonly, some ACF bypass the polyp stage in their carcinogenesis thus reinforcing the importance of their early detection and our understanding of their pathogenesis. Since ACF were first detected in carcinogen-treated mice, research efforts have focused on these microscopically visible lesions both in animal and human models. ACF show variable histological features, characterized by Kudo (20) and, therefore, can be grouped into differing categories by in vivo examination with high-magnification-chromoscopic-colonoscopy (HMCC). As expected, ACF are more frequently detected in distal animal and human colons coinciding with the geographic distribution of colorectal cancer (CRC). Various proteomic (Prot) markers may be altered within ACF suggesting possible prospective pathological changes. These markers include Calreticulin, Transgelin, Serotransferrin, Triphosphate isomerase and Carbonic anhydrase II. Other markers of importance include carcinoembryonic antigen (CEA), B-catenin, placental cadherin (P-cadherin), epithelial cadherin (E-cadherin), inducible nitric oxide synthase (iNOS), cyclooxygenase (COX-2) and P16INK4a. Genetic mutations of K-ras, B-Raf APC and p53 have been demonstrated in ACF as well as the epigenetic alterations of CpG island methylation. Genomic instabilities (GI), illustrated by a higher GI Index (GII), microsatellite instability (MSI), loss of heterozygosity (LOH) and defects in mismatch repair (MMR) systems, are also expressed. These transformations may lead to the identification of the earliest pathological features initiating colon tumorigenesis. In this review, the advances in ACF research as precursors of CRCs are highlighted.

Animals↗