SEARCH · Search PubMed

Results for “MME”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

SourceResearch connectionFrom yearThrough year

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 127 records · Page 7Linked to original sources

PubMed1988

The incorporation of monomethylethanolamine and dimethylethanolamine in fetal brain aggregating cell culture.

Fetal rat brain aggregating cell cultures were exposed to varying concentrations of [3H]monomethylethanolamine (MME) and [3H] dimethylethanolamine (DME). The rate of labeling of water-soluble compounds was more rapid and the amount of radioactivity present was greater than in the lipids. After a 72 hour incubation in the presence of millimolar concentrations of these nitrogenous bases, the major water-soluble products were the phosphorylated form of the bases. Little label was associated with the free bases or their cytidyl derivative. In the phospholipids, 97% of the radioactivity was recovered in phosphatidylmonomethylethanolamine (PMME) and 3% in phosphatidyldimethylethanolamine (PDME) or 95% in PDME and 5% in phosphatidylcholine (PC) after growth in presence of [3H]MME and [3H]DME respectively. The rate of formation of the radioactive products increased as function of the concentration of the nitrogenous base added up to 4 mM, the highest concentration employed. There was no significant difference in the pattern of labeling with cells grown in media devoid of methionine or choline. The turnover of the water-soluble metabolites was more rapid than in the phospholipids where an apparent half-life of 24 hours was calculated.

Animals↗

PubMed2006

High-resolution mapping of molecular events associated with immortalization, transformation, and progression to breast cancer in the MCF10 model.

BACKGROUND: A comprehensive and consistent picture of the genetic changes that underlie breast cancer initiation, development, and progression remains unresolved. The MCF10 series of cell lines represents many steps in that progression. We performed high resolution mapping of the MCF10 series of cell lines to identify specific gene targets to elucidate the molecular correlates of immortalization, development, and progression of breast cancer at the level of individual genes. DESIGN: We evaluated the initial untransformed outgrowths (MCF-10MS and MCF-10A) with six transformed cell lines with benign proliferations (MCF-10AT1, MCF-10AT1kcl2), carcinoma in situ (MCF-10CA1h cl13), and invasive carcinoma (MCF-10CA1h cl2, MCF-10CA1a cl1, MCF-10CA1d cl1). Losses and gains of loci at 112 unique human genome sites were interrogated using the multiplex ligation-dependent probe amplification assay (MLPA). RESULTS: Cytogenetic alterations in the four benign progenitors that persisted in the CIS and invasive cell lines corresponded to gains and losses of genes by MLPA. MCF-10MS had only normal gene copies. The untransformed MCF-10A had cytogenetic gain of 5q13-qter with corresponding gains of the IL3, IL4 and IL12B genes at 5q31-q33; gain of distal 19q12-qter was reflected in gains in KLK3 and BAX gene loci at 19q13-q13.4. The observed genic gain of cMYC at 8q24.12 was not indicated by cytogenetics. The apparently balanced t(3;9) component of the t(3;9)(p13;p22)t(3;5)(p26;q31) resulted in complete loss of the CDKN2A and CDKN2B genes at 9p21. Additional clonal cytogenetic changes in the DCIS cell line (MCF-10A1h cl13) involving chromosomes 1, 3 and 10 persisted in the invasive progeny, with gain of corresponding genes at 1p13 (BCAR2, BCAR3, NRAS, TGFB2), at 3p12-13 (IL12A), and 3q21-27 (MME, PIK3CA, BCL6). CONCLUSIONS: Our study adopted a comprehensive exploration of genetic changes using high resolution molecular probes applied to the MCF10 family of cell lines to identify individual genes in a continuum starting from normal breast epithelial cells and progressing through immortalization, transformation and invasive malignancy. Homozygous loss of CDKN2A and CDKN2B genes and gain of MYC were initiating immortalization events. Transformation and progression to malignancy event were marked by gains of IL13, VEGF, HRAS, TRAF2, and BCAS2, IL12A, and MME, respectively.

Adult↗

PubMed2006

Fatty acid composition of habitual omnivore and vegetarian diets.

High-fat diets are implicated in the onset of cardiovascular disease (CVD), cancer, and obesity. Large intakes of saturated and trans FA, together with low levels of PUFA, particularly long-chain (LC) omega-3 (n-3) PUFA, appear to have the greatest impact on the development of CVD. A high n-6:n-3 PUFA ratio is also considered a marker of elevated risk of CVD, though little accurate data on dietary intake is available. A new Australian food composition database that reports FA in foods to two decimal places was used to assess intakes of FA in four habitual dietary groups. Analysis using the database found correlations between the dietary intakes of LC n-3 PUFA and the plasma phospholipid LC n-3 PUFA concentrations of omnivore and vegetarian subjects. High meat-eaters (HME), who consumed large amounts of food generally, had significantly higher LC n-3 PUFA intakes (0.29 g/d) than moderate meat-eaters (MME) (0.14 g/d), whose intakes in turn were significantly higher than those of ovolacto-vegetarians or vegans (both 0.01 g/d). The saturated FA intake of MME subjects (typical of adult male Australians) was not different from ovolacto-vegetarian intakes, whereas n-6:n-3 intake ratios in vegetarians were significantly higher than in omnivores. Thus, accurate dietary and plasma FA analyses suggest that regular moderate consumption of meat and fish maintains a plasma FA profile possibly more conducive to good health.

Adult↗

PubMed1982

Left ventricular relaxation, mitral valve prolapse, and intracardiac conduction in myotonia atrophica: assessment by digitized echocardiography and noninvasive His bundle recording.

Myotonia atrophica, a neuromuscular disease marked by autosomal dominant transmission and delayed relaxation of skeletal muscle, has been associated with cardiac failure, conduction abnormality and mitral prolapse (MVP). In order to determine the relaxation rate of cardiac muscle, left ventricular (LV) size and function, and the presence of MVP, 30 patients with myotonia atrophica were studied using digitized M-mode echocardiography (MME). Intracardiac conduction intervals were determined by noninvasive His bundle recording (HBR) from surface electrodes using a high-resolution, R-wave triggered, signal averaging computer. Neurologically unaffected first-degree relatives of the patients with myotonia atrophica were also studied to determine if cardiac abnormalities may be present in the absence of neurologic manifestations of the disease. Peak normalized diastolic endocardial velocity in patients with myotonia atrophica (3.7 +/- 0.8 sec-1) did not differ from unaffected first-degree relatives (3.8 +/- 0.8 sec-1) or normal subjects (3.6 +/- 0.8 sec-1). Systolic LV function and LV dimensions on MME were normal in both groups. However, MVP was present in 7 of 24 (29%) of patients who could be evaluated, but not in unaffected first-degree relatives. Despite normal LV systolic and diastolic function, infranodal intracardiac conduction was prolonged in patients with myotonia atrophica (average HV interval 50 +/- 5 SD msec) but not in neurologically unaffected relatives (average HV interval 40 +/- 5 msec). Delay in proximal intracardiac conduction was also found in patients with myotonia atrophica (average PH interval 140 +/- 20 msec) but not in neurologically unaffected relatives (average PH interval 115 +/- 6 msec). Hence cardiac findings in myotonia atrophica include proximal and distal conduction delay by external HBR even in the absence of abnormality of the standard 12-lead ECG. There may also be an increased frequency of MVP; however, early diastolic relaxation of the LV is unimpaired, and cardiac manifestations of myotonia are not transmitted independently of neurologic abnormality.

Adolescent↗

PubMed2026

Comparison of Ketamine and Pregabalin on Postoperative Opioid Usage and Pain Management in Spinal Fusion: Systematic Review and Network Meta-analysis.

BACKGROUND CONTEXT: Spinal fusion is associated with substantial early postoperative pain and opioid exposure. Both ketamine and pregabalin are widely incorporated into Enhanced Recovery After Surgery (ERAS) protocols as opioid-sparing adjuncts. However, their comparative efficacy and safety in this specific setting remain uncertain. Our objective was to compare ketamine and pregabalin indirectly for early postoperative opioid consumption, pain, and adverse events in adults undergoing spinal fusion. METHODS: Pubmed, Embase, and Cochrane Trials were searched from inception through October 2025. Eligible studies were randomized trials enrolling adults undergoing instrumented spinal fusion, randomized to perioperative ketamine, pregabalin, or control, and reported extractable 24-hour opioid consumption or pain outcomes. Continuous outcomes were pooled as mean differences in MME or VAS units, and adverse events were reported descriptively. A connected treatment network was analyzed using random-effects models. Risk of bias (RoB) was assessed with the Cochrane RoB 2 tool. RESULTS: Thirteen trials (n=879) were included: ketamine (n=210), pregabalin (n=271), and control (n=398). Six trials contributed opioid data (3 ketamine, 3 pregabalin). Using pregabalin 150 mg as reference, ketamine was associated with lower 0-24-hour opioid use (MD -56.99 mg MME; 95% CI -99.56 to -14.43). Control (MD +21.31; 95% CI -1.05 to +43.66) and pregabalin 300 mg (MD -13.22; 95% CI -40.41 to +13.96) did not significantly differ from pregabalin 150 mg. Seven trials contributed 24-hour VAS data, with control being associated with higher pain versus pregabalin 150 mg (MD +0.84; 95% CI +0.01 to +1.66), while ketamine and pregabalin 300 mg were not k significantly different. Adverse events were generally infrequent and similar to control. CONCLUSIONS: Both ketamine and pregabalin provide early opioid sparing with comparable 24-hour analgesia. Ketamine showed a larger opioid-sparing point estimate, but indirect comparisons are imprecise. Adequately powered head-to-head trials with standardized protocols and adverse event reporting are needed.

Humans↗

PubMedDate not supplied

[Valpromide, Valproic acid and removal of small intestine in the treatment of a chronic depression: a case report].

Valpromide (VPD) is an antiepileptic drug, derivative of Valproic acid (VPA), used as a mood-stabilizer in bipolar disorder for 25 years in several European countries. VPD is also used as an augmentation strategy in refractory depression. Despite chemical similarity between VPA and VPD, the pharmacokinetics of the 2 drugs in humans are quite distinct. We report a case of a patient, suffering from a bipolar treatment resistant depression, who dramatically improved after substituting VPD to VPA in association with fluoxetine. Mme X, 68 years old, has been hospitalized in March 2001 for the treatment of a resistant depression (TRD). She was suffering from removal of small intestine with chronic diarrhoea after a suicidal attempt two years ago. She had a bipolar disorder treated with VPD (1,200 mg/d) since 1 year. She presented a major depressive episode according to DSM IV with various symptoms like depressed mood, hypersomnia and difficulty initiating sleep, diminished ability to concentrate and to think, markedly diminished pleasure in all activities and major anxiety. Mme X fulfilled TRD diagnosis after resistance to two adequate antidepressants trials from different classes (clomipramine 175 mg/day and venlafaxine 300 mg/day). The antidepressant treatment (venlafaxine) was interrupted and she has been receiving a SSRI (fluoxetine 20 mg/day) for 4 weeks. After four weeks, she had a partial remission with persistent sleep problems, mood lability and anxiety. The VPA blood concentration was very low: 27 mg/L (normal range: 50 to 100 mg/L) in spite of a high dosage: 1,200 mg/day. Pharmacokinetic analysis of VPD shown that VPD transformation to VPA usually done in the intestine, was reduced because of the removal of hail intestine. We substituted VPD by VPA. Valproate blood concentration returned to normal range, induced dramatic improvement of depression within three days. VPD is an amide derivative of valproic acid (valproate), biotransformed by hydrolysis to its corresponding valproic acid. VPD is a prodrug of VPA. VPD is absorbed after transformation in gastro-intestinal mucous membrane. The adequate dosage of VPD (Depamide, 300 mg) is 4 to 6 tablets in acute manic phases, 2 to 4 tablets in long term treatment, 1 to 3 tablets in depressive episode. The biodisponibility of VPD is around 100% 75 and 90% of VPD is linked with protein albumin. The daily dosage determined the blood concentration of the active form (VPA), but this relation isn't linear. The optimal blood concentration of VPA (Depakine) ranges between 50 and 100 mg/L. the free form of VPA is influenced by protein disorders such as of hypoalbuminemia and by presence of fat acids in food. This case report demonstrates at a clinical level that VPD and VPA are not equivalent for treating bipolar depression. This case also suggests that a deep investigation of the pharmacokinetic of psychotropic drugs can help clinicians to resolve clinical problems of treatment of depression.

Aged↗

PubMed2000

Differential regulation of type IV collagenases and metalloelastase in murine macrophages by the synthetic bacterial lipopeptide JBT 3002.

We determined whether the expression of matrix metalloproteinases (MMP) and tissue inhibitors of MMPs (TIMP) in murine macrophages is regulated by the novel synthetic bacterial lipopeptide JBT 3002. Multilamellar liposomes (MLV) encapsulating JBT 3002 (MLV-JBT 3002) stimulated the production of 72-kDa and 92-kDa (gelatinase A and B) type IV collagenase and inhibited the production of murine metalloelastase (MME) in a dose-dependent manner in murine peritoneal macrophages. MLV-JBT 3002 also induced production of TIMP-1. MLV-JBT 3002 did not induce collagenase production in tumor cells. Priming murine macrophages with interferon-gamma (IFN-gamma) inhibited JBT 3002-stimulated production of both MMP-9 and MMP-2 and further inhibited production of MME by a mechanism involving nitric oxide (NO). This conclusion is based on data showing that IFN-gamma failed to inhibit production of MMP in the presence of L-methyl arginine or in macrophages from inducible nitric oxide synthase knockout mice. These data suggest that JBT 3002 differentially regulates the production of various MMPs and TIMP in macrophages.

Adjuvants, Immunologic↗

PubMed2003

Muscle and temporomandibular joint forces associated with chincup loading predicted by numerical modeling.

Development of the components of the temporomandibular joint (TMJ) is thought to reflect joint loading. The aims of this project were to test 3 hypotheses: whether effective eminence morphology, masticatory muscle forces, and predicted TMJ forces during chincup loading of the mandible were consistent with the objectives of minimization of joint loads (MJL) or muscle effort (MME), or both. Regression relationships of MJL model-predicted versus measured eminence shapes in 9 subjects indicated a high degree of correlation (mean slope = 0.99, compared with perfect-match slope = 1.00). Model predictions of muscle output during chincup loading of the mandible were tested by comparison with data gathered in 6 subjects. Midsagittal plane chin loads were applied over a range of 60 degrees while bilateral masticatory muscle surface electromyography was quantified. The regression relationships of predicted versus measured masseter and anterior digastric muscle outputs indicated that model predictions were highly correlated (mean slope (masseter muscle) = 1.02; mean slope (digastric muscle) = 0.96). TMJ forces predicted by modeling showed intersubject differences of up to 34% for similar chincup loading conditions. Intrasubject variation in TMJ forces was as high as 57%, depending on chin load angle. The results demonstrated that TMJ eminence shape and masticatory muscle forces were consistent with objectives of both MJL and MME. Variation in TMJ forces depended on the subject and the direction of chincup loading.

Adolescent↗

PubMed1998

Sulfonamide-based hydroxamic acids as potent inhibitors of mouse macrophage metalloelastase.

The structural requirements of sulfonamide-based hydroxamic acid 1 for inhibition of macrophage metalloelastase (MME) were investigated. A short aliphatic group at the R2 position together with an aromatic group at the R3 position significantly improved the inhibitory activity. Compounds 32, 34, and 40 were the most potent inhibitors of MME with IC50 values between 5 and 6 nM.

Animals↗

PubMed2004

Frustrated beta-chloride elimination. Selective arene alkylation by alpha-chloronorbornene catalyzed by electrophilic metallocenium ion pairs.

Single-site polymerization catalysts generated in situ via activation of Cp*MMe(3) (Cp* = C(5)Me(5); M = Ti, Zr), (CGC)MMe(2) (CGC = C(5)Me(4)SiMe(2)NBu(t)(); M = Ti, Zr), and Cp(2)ZrMe(2) with Ph(3)C(+)B(C(6)F(5))(4)(-) catalyze alkylation of aromatic molecules (benzene, toluene) with alpha-chloronorbornene at room temperature, to regioselectively afford the 1:1 addition products exo-1-chloro-2-arylnorbornane (aryl = C(6)H(5) (1a), C(6)H(4)CH(3) (1b)) in good yields. Analogous deuterium-labeled products exo-1-chloro-2-aryl-d(n)-norbornane-7-d(1) (aryl-d(n) = C(6)D(5) (1a-d(6)), C(6)D(4)CD(3) (1b-d(8))) are obtained via catalytic arylation of alpha-chloronorbornene in either benzene-d(6) or toluene-d(8). Isolated ion-pair complexes such as (CGC)ZrMe(toluene)(+)B(C(6)F(5))(4)(-) and Cp(2)ThMe(+)B(C(6)F(5))(4)(-) also catalyze the reaction of alpha-chloronorbornene in toluene-d(8) to give 1b-d(8) in good yields, respectively. Small quantities of the corresponding bis(1-chloronorbornyl)aromatics 2 are also obtained from preparative-scale reactions. These reactions exhibit turnover frequencies exceeding 120 h(-1) (for the Cp*TiMe(3)/Ph(3)C(+)B(C(6)F(5))(4)(-)-catalyzed system), and chlorine-free products are not observed. Compounds 1 and 2 were characterized by (1)H, (2)H, (13)C, and 2D NMR, GC-MS, and elemental analysis. The aryl group exo-stereochemistry in 1a and 1b is established using (1)H-(1)H COSY, (1)H-(13)C HMBC, and (1)H-(1)H NOESY NMR, and is further corroborated by X-ray analysis of the product 1,4-bis(exo-1-chloro-2-norbornyl)benzene (2a). Control experiments and reactivity studies on each component step suggest a mechanism involving participitation of the metal electrophiles in the catalytic cycle.

Journal Article↗

PubMed2001

Alpha-1-antitrypsin and a broad spectrum metalloprotease inhibitor, RS113456, have similar acute anti-inflammatory effects.

There is increasing evidence that antiproteases are able to affect the inflammatory response. To further examine this question, we administered human alpha-1-antitrypsin (alpha1AT) or a synthetic metalloprotease inhibitor (RS113456) to C57 mice followed by a single intratracheal dose of quartz, a dust that evokes a marked, lasting, polymorphonuclear leukocyte (PMN) infiltrate. At 2 hours after dust administration, both antiproteases completely suppressed silica-induced PMN influx into the lung and macrophage inflammatory protein-2 (MIP-2)/monocyte chemotactic protein-1 (MCP-1) (neutrophil/macrophage chemoattractant) gene expression, partially suppressed nuclear transcription factor kappaB (NF-kappaB) translocation, and increased inhibitor of NF-kappaB (IkappaB) levels. By 24 hours, PMN influx and connective tissue breakdown measured as lavage desmosine or hydroxyproline were still at, or close to, control levels after antiprotease treatment, and increases in NF-kappaB translocation and MIP-2/MCP-1 gene expression were variably suppressed. At both time points, neither agent prevented silica-induced increases in amount of whole lung MIP-2 or MCP-1 protein, but both did prevent increases in whole lung intercellular adhesion molecule-1 (ICAM-1) at 24 hours. Inactivating the alpha1AT by oxidation to the point that it no longer possessed antiproteolytic properties did not affect its ability to suppress inflammation. Both antiproteases also prevented the silica-induced acute inflammatory response in mice with knocked out genes for macrophage metalloelastase (MME -/-), mice that develop inflammation, but not connective tissue breakdown, and the pattern of alpha1AT breakdown fragments was identical in control and MME -/- animals. These findings suggest that, in this model of acute PMN mediated inflammation, a serine protease inhibitor and a metalloprotease inhibitor have similar anti-inflammatory properties, that inflammation is not mediated by proteolysis with generation of chemotactic matrix fragments, and that classic antiproteolysis (complexing of protease to antiprotease) probably does not play a role in suppression of inflammation. The antiproteolytic effects of these agents do not seem to be mediated by protection of endogenous alpha1AT.

Animals↗

PubMed2006

Mechanisms of disease: Alloimmunization in renal diseases.

Graft rejection has long been considered the paradigm of renal diseases induced by alloimmunization, particularly alloimmunization directed against HLA antigens. Accumulating evidence indicates that non-HLA immunity also has an important role in clinical transplantation. Targets of alloimmunization include antigens of tubular basement membrane, tubular epithelial cells and endothelial cells. They can be polymorphic allovariants (as shown in the rat) or 'hidden' antigens exposed when the graft is damaged. Alloimmunization can also occur when a person genetically deficient in a renal protein (e.g. the alpha5 (IV) collagen chain in X-linked Alport's syndrome or nephrin in Finnish-type nephrotic syndrome) is transplanted to treat end-stage renal failure. The non-mutated protein in the donor kidney is recognized as a foreign antigen, and the resulting alloimmune response can damage the graft. We have demonstrated that alloimmunity can also affect the native kidney. We have characterized a novel fetomaternal disease in which a genetic defect in the MME gene encoding neutral endopeptidase (NEP) in the mother leads to the development of membranous nephropathy in her fetus (maternal anti-NEP antibodies bind to NEP on fetal podocytes). Our findings raise the possibility that mutations or genetic polyporphisms in MME or other genes expressed by the podocyte are involved in alloimmune-mediated development of membranous nephropathy after kidney or bone marrow transplantation.

Humans↗

PubMed2026

Liposomal bupivacaine versus ropivacaine for surgical site infiltration in lumbar fusion: a prospective randomized controlled trial.

INTRODUCTION: Effective postoperative pain control after lumbar spine surgery remains challenging, and excessive opioid use is associated with adverse outcomes. Evidence comparing liposomal bupivacaine (LB) with conventional long-acting local anesthetics in spine surgery is limited. PATIENTS AND METHODS: In this single-center, prospective, randomized, patient- and outcome assessor-blinded trial, adult undergoing one- or two-level posterior lumbar decompression and fusion were assigned (1:1) to surgical site infiltration with either LB (266 mg) plus 25 mg plain bupivacaine (LB group) or ropivacaine (R group). The primary outcome was 72 h cumulative opioid consumption (morphine milligram equivalents, MME). Secondary outcomes included time-profile opioid consumption, pain scores, rescue analgesia, safety, and functional recovery. RESULTS: A total of 202 patients were included in the modified intention-to-treat analysis. Cumulative MME within 72 h was significantly lower in the LB group compared with the R group [43.0 (37.0, 58.0) mg vs. 58.0 (46.0, 73.0) mg], corresponding to a 22% relative reduction (GMR 0.78, 95% CI 0.71-0.85; p < 0.001). The reduction was most pronounced during 8-24 h and 24-48 h postoperatively. Overall pain scores at rest and with movement, as well as 72-h pain AUC, were lower in the LB group. No significant between-group differences were observed in rescue analgesia, adverse events and functional recovery. CONCLUSION: In patients undergoing one- or two-level posterior lumbar decompression and fusion, surgical site infiltration with an LB-based combined regimen, compared with ropivacaine monotherapy, reduced 72-h opioid consumption and cumulative postoperative pain burden without an observed increase in adverse events or impairment of early functional recovery.

Humans↗

PubMed1998

Angiostatin-mediated suppression of cancer metastases by primary neoplasms engineered to produce granulocyte/macrophage colony-stimulating factor.

We determined whether tumor cells consistently generating granulocyte/macrophage colony- stimulating factor (GM-CSF) can recruit and activate macrophages to generate angiostatin and, hence, inhibit the growth of distant metastasis. Two murine melanoma lines, B16-F10 (syngeneic to C57BL/6 mice) and K-1735 (syngeneic to C3H/HeN mice), were engineered to produce GM-CSF. High GM-CSF (>1 ng/10(6) cells)- and low GM-CSF (<10 pg/10(6) cells)-producing clones were identified. Parental, low, and high GM-CSF-producing cells were injected subcutaneously into syngeneic and into nude mice. Parental and low-producing cells produced rapidly growing tumors, whereas the high-producing cells produced slow-growing tumors. Macrophage density inversely correlated with tumorigenicity and directly correlated with steady state levels of macrophage metalloelastase (MME) mRNA. B16 and K-1735 subcutaneous (s.c.) tumors producing high levels of GM-CSF significantly suppressed lung metastasis of 3LL, UV-2237 fibrosarcoma, K-1735 M2, and B16-F10 cells, but parental or low-producing tumors did not. The level of angiostatin in the serum directly correlated with the production of GM-CSF by the s.c. tumors. Macrophages incubated with medium conditioned by GM-CSF- producing B16 or K-1735 cells had higher MME activity and generated fourfold more angiostatin than control counterparts. These data provide direct evidence that GM-CSF released from a primary tumor can upregulate angiostatin production and suppress growth of metastases.

Angiostatins↗

PubMed1988

Saccharomyces cerevisiae cho2 mutants are deficient in phospholipid methylation and cross-pathway regulation of inositol synthesis.

Five allelic Saccharomyces cerevisiae mutants deficient in the methylation of phosphatidylethanolamine (PE) have been isolated, using two different screening techniques. Biochemical analysis suggested that these mutants define a locus, designated CHO2, that may encode a methyltransferase. Membranes of cho2 mutant cells grown in defined medium contain approximately 10% phosphatidylcholine (PC) and 40-50% PE as compared to wild-type levels of 40-45% PC and 15-20% PE. In spite of this greatly altered phospholipid composition, cho2 mutant cells are viable in defined medium and are not auxotrophic for choline or other phospholipid precursors such as monomethylethanolamine (MME). However, analysis of yeast strains carrying more than one mutation affecting phospholipid biosynthesis indicated that some level of methylated phospholipid is essential for viability. The cho2 locus was shown by tetrad analysis to be unlinked to other loci affecting phospholipid synthesis. Interestingly, cho2 mutants and other mutant strains that produce reduced levels of methylated phospholipids are unable to properly repress synthesis of the cytoplasmic enzyme inositol-1-phosphate synthase. This enzyme was previously shown to be regulated at the level of mRNA abundance in response to inositol and choline in the growth medium. We cloned the CHO2 gene on a 3.6-kb genomic DNA fragment and created a null allele of cho2 by disrupting the CHO2 gene in vivo. The cho2 disruptant, like all other cho2 mutants, is viable, exhibits altered regulation of inositol biosynthesis and is not auxotrophic for choline or MME.

Choline↗

PubMed1989

Mutations in the Saccharomyces cerevisiae opi3 gene: effects on phospholipid methylation, growth and cross-pathway regulation of inositol synthesis.

We report the isolation of two new opi3 mutants by EMS mutagenesis, and construction of an insertion allele in vitro using the cloned gene. We have demonstrated that the opi3 mutations cause a deficiency in the two terminal phospholipid N-methyltransferase (PLMT) activities required for the de novo synthesis of PC (phosphatidylcholine). The opi3 mutants, under certain growth conditions, produce membrane virtually devoid of PC although, surprisingly, none of the mutants displays a strict auxotrophic requirement for choline. Although the opi3 mutants grow without supplements, we have shown that the atypical membrane affects the ability of the mutant strains to initiate log phase growth and to sustain viability at stationary phase. The commencement of log phase growth is enhanced by addition of choline or to a lesser extent DME (dimethylethanolamine), and retarded by addition of MME (monomethylethanolamine). The mutant cells lose viability at the stationary phase of the cell cycle in the absence of DME or choline, and are also temperature sensitive for growth at 37 degrees especially in media containing MME. These growth defects have been correlated to the presence of specific phospholipids in the membrane. The opi3 growth defects are suppressed by an unusual mutation in the phospholipid methylation pathway that perturbs the N-methyltransferase (PEMT) activity immediately preceding the reactions affected by the opi3 lesion. We believe this mutation, cho2-S, alters the substrate specificity of the PEMT. A secondary effect of opi3 mutations is disruption of the cross pathway regulation of the synthesis of the PI (phosphatidylinositol) precursor inositol. Synthesis of inositol is controlled through regulation of the INO1 gene which encodes inositol-1-phosphate synthase. This highly regulated gene is expressed constitutively in opi3 mutants. We have used the opi3 strains to demonstrate that synthesis of either PC or PD (phosphatidyldimethylethanolamine) will restore normal regulation of the INO1 gene.

Alleles↗

PubMed2007

A novel ex vivo skin model to study the susceptibility of the dermatophyte Trichophyton rubrum to photodynamic treatment in different growth phases.

BACKGROUND: Dermatophytes are fungi that can cause infections of skin, hair and nails because of their ability to feed on keratin. Superficial mycoses are among the most prevalent infectious diseases worldwide. Two important restrictions of current therapeutic options are the recurrence of the infection and prolonged treatment. This is especially true for infections caused by Trichophyton rubrum, a widely distributed dermatophyte. The application of photosensitizers for treatment of fungal infections is, within the field of photodynamic treatment (PDT), relatively new. Recently, we demonstrated that the porphyrins 5,10,15-tris(4-methylpyridinium)-20-phenyl-[21H,23H]-porphine trichloride (Sylsens B) and deuteroporphyrin monomethylester (DP mme) were excellent photosensitizers towards T. rubrum when using red light. OBJECTIVES AND METHODS: To evaluate the photodynamic effectiveness of the porphyrins in a situation that mimics the clinical situation, we developed an ex vivo model using human stratum corneum. This model offers the possibility of applying PDT at different time points during the germination and subsequent development of T. rubrum microconidia. The model was used for two different incubation media, Dulbecco's modified Eagle medium (DMEM) and distilled water. RESULTS AND CONCLUSIONS: We demonstrated that the PDT susceptibility of T. rubrum depended on the time of PDT application after spore inoculation. A decrease in susceptibility was observed with increasing time of PDT application for both photosensitizers in DMEM. Changing the incubation medium to distilled water resulted in an increased fungicidal effect for Sylsens B and in a decreased effect for DP mme. We conclude that T. rubrum is susceptible to PDT in a situation that mimics the clinical situation. The fungicidal effect of PDT on fungal spores is of particular importance.

Culture Media↗

PubMed2005

DNA analysis with multiplex microarray-enhanced PCR.

We have developed a highly sensitive method for DNA analysis on 3D gel element microarrays, a technique we call multiplex microarray-enhanced PCR (MME-PCR). Two amplification strategies are carried out simultaneously in the reaction chamber: on or within gel elements, and in bulk solution over the gel element array. MME-PCR is initiated by multiple complex primers containing gene-specific, forward and reverse, sequences appended to the 3' end of a universal amplification primer. The complex primer pair is covalently tethered through its 5' end to the polyacryl- amide backbone. In the bulk solution above the gel element array, a single pair of unattached universal primers simultaneously directs pseudo-monoplex PCR of all targets according to normal solution-phase PCR. The presence of a single universal PCR primer pair in solution accelerates amplification within gel elements and eliminates the problem of primer interference that is common to conventional multiplex PCR. We show 10(6)-fold amplification of targeted DNA after 50 cycles with average amplification efficiency 1.34 per cycle, and demonstrate specific on-chip amplification of six genes in Bacillus subtilis. All six genes were detected at 4.5 pg of bacterial genomic DNA (equivalent to 10(3) genomes) in 60 independent amplification reactions performed simultaneously in single reaction chamber.

DNA↗