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2.
Sci Total Environ ; 654: 841-849, 2019 Mar 01.
Article in English | MEDLINE | ID: mdl-30448673

ABSTRACT

Agriculture contributes 18% of India's greenhouse gas (GHG) emissions. Yet, little is known about the energy requirements of individual crops, making it difficult to link nutrition-enhancing dietary changes to energy consumption and climate change. We estimate the energy and CO2 intensity of food grains (rice, wheat, sorghum, maize, pearl millet and finger millet) taking into account their irrigation requirements, water source, dependence on groundwater, yields, fertilizer and machinery inputs. Rice is the most energy-intensive cereal, while millets are the least. Total energy use contributes 16% of GHG emissions for rice, due to its high methane emissions, and 56% for wheat. Fertilizer production and use dominates GHG emissions from all crops, contributing 52% of GHGs from cereals. Energy intensities vary by up to a factor of four across the country, due to varying water requirements, irrigation sources and groundwater table depths. The results suggest that replacing rice with other cereals has the potential to reduce energy consumption and GHGs, though the spatial variation of production shifts would influence the extent of this reduction and the possible trade-offs with total production.


Subject(s)
Air Pollutants/analysis , Crop Production , Edible Grain/growth & development , Greenhouse Gases/analysis , Methane/analysis , Nitrous Oxide/analysis , Agricultural Irrigation , Fertilizers/analysis , Groundwater/analysis , India
3.
J Vet Intern Med ; 29(1): 104-12, 2015 Jan.
Article in English | MEDLINE | ID: mdl-25537303

ABSTRACT

BACKGROUND: Acid suppressant drugs are a mainstay of treatment for cats with gastrointestinal erosion and ulceration. However, clinical studies have not been performed to compare the efficacy of commonly PO administered acid suppressants in cats. HYPOTHESIS/OBJECTIVES: To compare the effect of PO administered famotidine, fractionated omeprazole tablet (fOT), and omeprazole reformulated paste (ORP) on intragastric pH in cats. We hypothesized that both omeprazole formulations would be superior to famotidine and placebo. ANIMALS: Six healthy adult DSH colony cats. METHODS: Utilizing a randomized, 4-way crossover design, cats received 0.88-1.26 mg/kg PO q12h fOT, ORP, famotidine, and placebo (lactose capsules). Intragastric pH monitoring was used to continuously record intragastric pH for 96 hours beginning on day 4 of treatment. Plasma omeprazole concentrations at steady state (day 7) were determined by high performance liquid chromatography (HPLC) with ultraviolet detection. Mean percentage time that intragastric pH was ≥ 3 and ≥ 4 were compared among groups using ANOVA with a posthoc Tukey-Kramer test (α = 0.05). RESULTS: The mean percentage time ± SD that intragastric pH was ≥ 3 was 68.4 ± 35.0% for fOT, 73.9 ± 23.2% for ORP, 42.8 ± 18.6% for famotidine, and 16.0 ± 14.2% for placebo. Mean ± SD plasma omeprazole concentrations were similar in cats receiving fOT compared to those receiving ORP and in a range associated with acid suppression reported in other studies. CONCLUSIONS AND CLINICAL IMPORTANCE: These results suggest that both omeprazole formulations provide superior acid suppression in cats compared to famotidine or placebo. Fractionated enteric-coated OT is an effective acid suppressant despite disruption of the enteric coating.


Subject(s)
Antacids/pharmacology , Cats/physiology , Famotidine/pharmacology , Omeprazole/pharmacology , Stomach/physiology , Animals , Antacids/administration & dosage , Antacids/pharmacokinetics , Anti-Ulcer Agents/administration & dosage , Anti-Ulcer Agents/pharmacology , Area Under Curve , Cross-Over Studies , Half-Life , Hydrogen-Ion Concentration , Monitoring, Physiologic/instrumentation , Monitoring, Physiologic/veterinary , Ointments , Omeprazole/administration & dosage , Omeprazole/pharmacokinetics , Tablets
4.
Clin Microbiol Infect ; 16(8): 1142-8, 2010 Aug.
Article in English | MEDLINE | ID: mdl-19832715

ABSTRACT

To assess the role of non-tuberculous mycobacteria (NTM) as a cause of tuberculosis-like diseases in Zambia, 167 chronically ill patients, hospitalized in three rural hospitals in Katete, Sesheke and Chilonga, were examined by microscopy and liquid culture for the presence of NTM. The percentages of patients with a positive culture for Mycobacterium tuberculosis complex were similar in the three geographical locations (19-25%). In contrast, the percentage of NTM ranged from 78% in Katete and 65% in Sesheke to 21% in Chilonga. Furthermore, the distribution of NTM species was different at the three geographical sites. In seven patients, true NTM-associated disease was suspected: five with Mycobacterium lentiflavum and two with Mycobacterium intracellulare. Analysis of possible risk factors indicated that the OR for NTM culture-positive sputum was significantly higher for patients living in Katete and Sesheke. Female gender and chest X-ray appearances of tuberculosis were independently associated with NTM culture-positive sputum. NTM colonization and disease in hospitalized, chronically ill patients in rural Zambia appear to be common.


Subject(s)
Mycobacterium Infections, Nontuberculous/epidemiology , Mycobacterium Infections, Nontuberculous/microbiology , Nontuberculous Mycobacteria/classification , Nontuberculous Mycobacteria/isolation & purification , Adolescent , Adult , Aged , Child , Female , Humans , Male , Middle Aged , Mycobacterium tuberculosis/isolation & purification , Pilot Projects , Prevalence , Radiography, Thoracic , Risk Factors , Rural Population , Tuberculosis/epidemiology , Tuberculosis/microbiology , Young Adult , Zambia/epidemiology
5.
Afr J Paediatr Surg ; 6(1): 3-6, 2009.
Article in English | MEDLINE | ID: mdl-19661656

ABSTRACT

BACKGROUND: In African settings the treatment results of Burkitt's lymphoma (BL) seem to be less favourable compared with Western settings. The aim of this retrospective study was to analyse some factors that affect the treatment of BL. PATIENTS AND METHODS: Over a 16 year period, data were extracted of 80 patients. RESULTS: Complete remission 5%, very good partial response 35%, partial response 16%, no response 10%, data missing 34%. Of all patients, 56% did have a positive response to treatment. However, 51% of this subgroup of patients did not finish treatment. There was no difference in completion of treatment between patients living in Katete district finishing treatment vs. living outside Katete district (respectively 25% vs. 32%, P = 0.7148). CONCLUSION: There is potential for higher cure rates for BL in tropical settings if full effort is put into compliance since a majority of patients, even while having a good prognosis, abandon treatment. Large distance to hospital makes no difference in completing the chemotherapy course.


Subject(s)
Antineoplastic Agents/therapeutic use , Burkitt Lymphoma/drug therapy , Health Services Accessibility , Patient Compliance , Adolescent , Adult , Child , Child, Preschool , Female , Humans , Male , Middle Aged , Retrospective Studies , Rural Population , Treatment Outcome , Zambia
6.
Article in English | AIM (Africa) | ID: biblio-1257510

ABSTRACT

Background : In African settings the treatment results of Burkitt's lymphoma (BL) seem to be less favourable compared with Western settings. The aim of this retrospective study was to analyse some factors that affect the treatment of BL. Patients and Methods : Over a 16 year period; data were extracted of 80 patients. Results : Complete remission 5; very good partial response 35; partial response 16; no response 10; data missing 34. Of all patients; 56did have a positive response to treatment. However; 51of this subgroup of patients did not finish treatment. There was no difference in completion of treatment between patients living in Katete district finishing treatment vs. living outside Katete district (respectively 25vs. 32; P = 0.7148). Conclusion: There is potential for higher cure rates for BL in tropical settings if full effort is put into compliance since a majority of patients; even while having a good prognosis; abandon treatment. Large distance to hospital makes no difference in completing the chemotherapy course


Subject(s)
Antineoplastic Agents/therapeutic use , Burkitt Lymphoma/drug therapy , Health Services Accessibility , Patient Compliance , Retrospective Studies , Rural Population , Treatment Outcome , Zambia
7.
Int J Tuberc Lung Dis ; 12(12): 1485-8, 2008 Dec.
Article in English | MEDLINE | ID: mdl-19017462

ABSTRACT

Polymorphism in various genes that may influence susceptibility to tuberculosis (TB) was examined in 46 TB patients and 119 healthy tuberculin-positive controls in Zambia. The odds of having TB was 2.8-fold higher in carriers of the -2518 AG single-nucleotide polymorphism in the promoter region of the CC-chemokine ligand 2 than in those carrying the homozygous genotype AA (95%CI 1.3-5.5).


Subject(s)
Chemokine CCL2/genetics , Genetic Predisposition to Disease , Tuberculosis/genetics , Adult , Female , Humans , Male , Polymorphism, Single Nucleotide , Zambia
8.
Genome ; 49(7): 825-39, 2006 Jul.
Article in English | MEDLINE | ID: mdl-16936791

ABSTRACT

The nucleolus organizer region (NOR) and 5S ribosomal RNA (rRNA) genes are valuable as chromosome landmarks and in evolutionary studies. The NOR intergenic spacers (IGS) and 5S rRNA nontranscribed spacers (NTS) were PCR-amplified and sequenced from 5 cultivars of the Andean grain crop quinoa (Chenopodium quinoa Willd., 2n = 4x = 36) and a related wild ancestor (C. berlandieri Moq. subsp. zschackei (Murr) A. Zobel, 2n = 4x = 36). Length heterogeneity observed in the IGS resulted from copy number difference in subrepeat elements, small re arrangements, and species-specific indels, though the general sequence composition of the 2 species was highly similar. Fifteen of the 41 sequence polymorphisms identified among the C. quinoa lines were synapomorphic and clearly differentiated the highland and lowland ecotypes. Analysis of the NTS sequences revealed 2 basic NTS sequence classes that likely originated from the 2 allopolyploid subgenomes of C. quinoa. Fluorescence in situ hybridization (FISH) analysis showed that C. quinoa possesses an interstitial and a terminal pair of 5S rRNA loci and only 1 pair of NOR, suggesting a reduction in the number of rRNA loci during the evolution of this species. C. berlandieri exhibited variation in both NOR and 5S rRNA loci without changes in ploidy.


Subject(s)
Chenopodium/genetics , DNA, Ribosomal Spacer/genetics , Genes, Plant , Polymorphism, Genetic , RNA, Ribosomal/genetics , Base Sequence , Chenopodium quinoa/genetics , Evolution, Molecular , In Situ Hybridization, Fluorescence , Molecular Sequence Data , Nucleolus Organizer Region , Phylogeny , RNA, Ribosomal, 5S/genetics , Sequence Alignment
9.
J Bone Joint Surg Br ; 88(6): 799-806, 2006 Jun.
Article in English | MEDLINE | ID: mdl-16720777

ABSTRACT

We reviewed retrospectively 45 patients (46 procedures) with bladder exstrophy treated by bilateral oblique pelvic osteotomy in conjunction with genitourinary repair. The operative technique and post-operative management with or without external fixation are described. A total of 21 patients attended a special follow-up clinic and 24 were interviewed by telephone. The mean follow-up time was 57 months (24 to 108). Of the 45 patients, 42 reported no pain or functional disability, although six had a waddling gait and two had marked external rotation of the hip. Complications included three cases of infection and loosening of the external fixator requiring early removal with no deleterious effect. Mid-line closure failed in one neonate managed in plaster. This patient underwent a successful revision procedure several months later using repeat osteotomies and external fixation. The percentage pubic approximation was measured on anteroposterior radiographs pre-operatively, post-operatively and at final follow-up. The mean approximation was 37% (12% to 76%). It varied markedly with age and was better when external fixation was used. The wide range reflects the inability of the anterior segment to develop naturally in spite of close approximation at operation. We conclude that bilateral oblique pelvic osteotomy with or without external fixation is useful in the management of difficult primary closure in bladder exstrophy, failed primary closure and secondary reconstruction.


Subject(s)
Bladder Exstrophy/surgery , Epispadias/surgery , Osteotomy/methods , Pelvis/surgery , Age Factors , Child , Child, Preschool , Cloaca/abnormalities , Cloaca/surgery , External Fixators , Female , Gait/physiology , Humans , Image Processing, Computer-Assisted , Infant , Male , Osteotomy/instrumentation , Pubic Bone/surgery , Reoperation , Retrospective Studies , Treatment Outcome , Urologic Surgical Procedures/methods
10.
Theor Appl Genet ; 112(8): 1593-600, 2006 May.
Article in English | MEDLINE | ID: mdl-16586115

ABSTRACT

Quinoa (Chenopodium quinoa Willd.) is adapted to the harsh environments of the Andean Altiplano region. Its seeds have a well-balanced amino acid composition and exceptionally high protein content with respect to human nutrition. Quinoa grain is a staple in the diet of some of the most impoverished people in the world. The plant is an allotetraploid displaying disomic inheritance (2n=4x=36) with a di-haploid genome of 967 Mbp (megabase pair), or 2C=2.01 pg. We constructed two quinoa BAC libraries using BamHI (26,880 clones) and EcoRI (48,000 clones) restriction endonucleases. Cloned inserts in the BamHI library average 113 kb (kilobase) with approximately 2% of the clones lacking inserts, whereas cloned inserts in the EcoRI library average 130 kb and approximately 1% lack inserts. Three plastid genes used as probes of high-density arrayed blots of 73,728 BACs identified approximately 2.8% of the clones as containing plastid DNA inserts. We estimate that the combined quinoa libraries represent at least 9.0 di-haploid nuclear genome equivalents. An average of 12.2 positive clones per probe were identified with 13 quinoa single-copy ESTs as probes of the high-density arrayed blots, suggesting that the estimate of 9.0x coverage of the genome is conservative. Utility of the BAC libraries for gene identification was demonstrated by probing the library with a partial sequence of the 11S globulin seed storage protein gene and identifying multiple positive clones. The presence of the 11S globulin gene in four of the clones was verified by direct comparison with quinoa genomic DNA on a Southern blot. Besides serving as a useful tool for gene identification, the quinoa BAC libraries will be an important resource for physical mapping of the quinoa genome.


Subject(s)
Chenopodium quinoa/genetics , Chromosomes, Artificial, Bacterial , Gene Library , Genes, Plant , Seeds/genetics , Cell Nucleus/chemistry , DNA, Plant/analysis , DNA, Plant/isolation & purification , Plant Proteins/chemistry , Plant Proteins/genetics
11.
Apoptosis ; 9(6): 807-14, 2004 Nov.
Article in English | MEDLINE | ID: mdl-15505423

ABSTRACT

Corpora lutea (CL) were isolated from one rabbit ovary on days 4, 8, 16 (peak luteal function), 28 (functional regression) and 30 of pregnancy and processed for biochemical analysis of DNA integrity. Analysis of DNA integrity revealed the presence of oligonucleosomal fragments in day 28 and day 30 CL but not in day 16 CL. The extent of low molecular weight (<15 kb) DNA labeling was 6.6 +/- 0.84 fold higher in day 30 as compared to day 16 CL (mean +/- SEM; n = 4, P < 0.01). In a second series of experiments, healthy CL collected from day 16 pregnant rabbits were incubated for 2 h in the absence or presence of 250 microg/ml of placental extract (PE) obtained from day 16 and/or day 30 placentas. Analysis of DNA integrity revealed that extensive apoptosis occurred in CL incubated in medium alone and in medium containing day 30 PE. In contrast, day 16 PE significantly suppressed apoptosis vs control (70 +/- 4%). In a third series of experiments, expression of mRNA for bcl-x and bax was measured by Northern analysis of CL treated without and with day 16 PE using cRNA probes for bcl-x and bax developed in our laboratory by RT-PCR. Treatment with PE significantly reduced bax mRNA levels but did not change bcl-x mRNA levels. These studies provide evidence that functional luteolysis in the pregnant rabbit CL is correlated with the occurrence of apoptosis. The data suggest that a factor(s) derived from the placenta may be responsible for the prolongation of CL life span during pregnancy by its ability to alter the bax:bcl -x rheostat and suppress apoptosis.


Subject(s)
Apoptosis , Corpus Luteum/physiology , Luteolysis , Placenta/chemistry , Pregnancy/physiology , Animals , Cell Survival , Female , Organ Culture Techniques , Progesterone/analysis , Proto-Oncogene Proteins c-bcl-2/genetics , Proto-Oncogene Proteins c-bcl-2/metabolism , RNA, Messenger/analysis , Rabbits , Radioimmunoassay , Time Factors , bcl-2-Associated X Protein , bcl-X Protein
12.
Radiat Prot Dosimetry ; 111(1): 51-4, 2004.
Article in English | MEDLINE | ID: mdl-15367768

ABSTRACT

Radon gas has been shown to cause an increased incidence of lung cancer. In affected areas, levels in the overground workplace can be sufficiently high to be a health risk and remediation is required. In the UK, the workplace Action Level is 400 Bq m(-3). The variation of radon levels in the workplace was studied both before and after remediation. In most rooms, remediation resulted in a greater reduction at night than during the working day. The dose reduction, and therefore the health benefit, to workers is less than that predicted by the drop in radon averaged over 24 h. In order to obtain a health benefit to 75% of workers in our series, the 24 h average radon level in each room must be reduced to <225 Bq m(-3). It is recommended that UK Regulatory Agencies adopt a post-remediation workplace Action Level of 225 Bq m(-3).


Subject(s)
Air Pollutants, Occupational/analysis , Air Pollutants, Radioactive/analysis , Air Pollution, Indoor/analysis , Environmental Monitoring/statistics & numerical data , Occupational Exposure/analysis , Radiometry/statistics & numerical data , Radon/analysis , Air Pollutants, Occupational/standards , Air Pollutants, Radioactive/standards , England , Humans , Maximum Allowable Concentration , Meteorological Concepts , Occupational Exposure/prevention & control , Occupational Exposure/standards , Pilot Projects , Time Factors , Workplace
14.
Biochem Soc Trans ; 29(Pt 6): 860-3, 2001 Nov.
Article in English | MEDLINE | ID: mdl-11709088

ABSTRACT

Protein kinase cascades feature in many signal transduction pathways. For those discussed here, a single upstream protein kinase appears to be responsible for the control of multiple downstream targets. So how is specificity introduced into these events? For the downstream kinases (substrates) described here, it would appear that specificity is determined by substrate-directed events that are permissive for phosphorylation. There are also distinctions relating to the turnover of these phosphorylations providing a further element of specificity.


Subject(s)
Cyclic AMP-Dependent Protein Kinases/metabolism , Protein Kinase C/metabolism , Allosteric Site , Animals , Cyclic AMP-Dependent Protein Kinases/chemistry , Models, Biological , Phosphatidylinositol 3-Kinases/metabolism , Phosphoinositide-3 Kinase Inhibitors , Phosphorylation , Protein Structure, Tertiary , Signal Transduction
15.
Reprod Fertil Dev ; 13(5-6): 395-403, 2001.
Article in English | MEDLINE | ID: mdl-11833936

ABSTRACT

Apoptosis is a morphologically defined type of cell death initiated by various stimuli that results in the activation of caspases (cysteine-containing aspartate-specific proteases). In the present study, it was determined that caspases are present during, and play a role in, corpus luteum (CL) apoptosis in vitro. Pseudopregnancy was induced in rabbits with 100 IU human chorionic gonadotrophin. On Day 11 of pseudopregnancy, CL were isolated and cultured for 0, 2, 4, 6, and 8 h in the absence of trophic support to induce spontaneous apoptosis. Total RNA was extracted and analysed for caspase-I expression by Northern blot analysis. The results demonstrated caspase-I expression from 4 h. In the second part of the study, CL were incubated without trophic support for 4 h with increasing concentrations of three general caspase inhibitors, sodium aurothiomalate (SAM), iodoacetic acid (IAA) and N-tosyl-L-phenylalanylchloromethylketone (TPCK), and two specific caspase inhibitors, N-acetyl (Ac)-Tyr-Val-Ala-Asp (YVAD)-chloromethylketone (CMK) (Ac-YVAD-CMK) and Ac-Asp-Glu-Val-Asp (DEVD)-aldehyde (CHO) (Ac-DEVD-CHO). At completion, DNA was isolated and integrity assessed. Treatment of CL with SAM, IAA or Ac-DEVD-CHO effectively suppressed apoptotic DNA fragmentation. The final component of the study was to examine caspase-3 protein expression. Western blot analysis revealed a significant increase in caspase-3 expression over the experimental time-course. The results of the present study clearly demonstrate a time-dependent link between the caspases, specifically caspase-3 and spontaneous apoptosis in the rabbit CL.


Subject(s)
Apoptosis/drug effects , Caspase Inhibitors , Corpus Luteum/cytology , Cysteine Proteinase Inhibitors/pharmacology , Amino Acid Chloromethyl Ketones/pharmacology , Animals , Blotting, Western , Caspase 1/genetics , Caspase 3 , Caspases/genetics , Chorionic Gonadotropin/pharmacology , Corpus Luteum/drug effects , Corpus Luteum/enzymology , DNA Fragmentation/drug effects , Female , Gene Expression , Gold Sodium Thiomalate/pharmacology , Humans , Iodoacetic Acid/pharmacology , Kinetics , Oligopeptides/pharmacology , Organ Culture Techniques , Phenotype , Pseudopregnancy , RNA, Messenger/analysis , Rabbits , Tosylphenylalanyl Chloromethyl Ketone/pharmacology
16.
Dig Dis Sci ; 45(2): 298-305, 2000 Feb.
Article in English | MEDLINE | ID: mdl-10711442

ABSTRACT

Guanylyl cyclase C is a sensitive and specific biomarker for metastatic colorectal cancer. A variant of the guanylyl cyclase C transcript was identified that possesses a 142-bp deletion at the 3' end of exon 1 reflecting alternative splicing of mRNA, introducing a shift in the open reading frame that prevents translation of a guanylyl cyclase C-related product. This variant was identified in human intestine and colon carcinomas, but not in extraintestinal tissues or tumors. These studies demonstrate that GCC and the splice variant contribute to the pool of GCC transcripts detected by RT-PCR in human tissues. They indicate that primers for RT-PCR that amplify regions downstream from the deletion are required to assess the full complement of GCC transcripts (GCC + GCC(var)) in human tissues and body fluids for staging and postoperative surveillance of patients with colorectal cancer.


Subject(s)
Adenocarcinoma/genetics , Colonic Neoplasms/genetics , Colorectal Neoplasms/genetics , Gene Expression Regulation, Neoplastic , Guanylate Cyclase/genetics , Receptors, Peptide/genetics , Transcription, Genetic , Antisense Elements (Genetics) , Colonic Neoplasms/pathology , Colorectal Neoplasms/pathology , Humans , RNA Splicing , Receptors, Enterotoxin , Receptors, Guanylate Cyclase-Coupled , Reverse Transcriptase Polymerase Chain Reaction
18.
Biochemistry ; 38(20): 6441-8, 1999 May 18.
Article in English | MEDLINE | ID: mdl-10350462

ABSTRACT

Calcium (Ca2+) and cyclic GMP (cGMP) subserve antagonistic functions that are reflected in their coordinated reciprocal regulation in physiological systems. However, molecular mechanisms by which Ca2+ regulates cGMP-dependent signaling remain incompletely defined. In this study, the inhibition of recombinant nitric oxide (NO)-stimulated soluble guanylyl cyclase (SGC) by Ca2+ was demonstrated. The alpha- and beta-subunits of recombinant rat SGC were heterologously coexpressed in HEK 293 cells which do not express NO synthase, whose Ca2+-stimulated activity can confound the effects of that cation on SGC. Ca2+ inhibited basal and NO-stimulated SGC in a concentration- and guanine nucleotide-dependent fashion. This cation inhibited SGC in crude cell extracts and immunopurified preparations. Ca2+ lowered both the Vmax and Km of SGC via an uncompetitive mechanism through direct interaction with the enzyme. In intact HEK 293 cells, increases in the intracellular Ca2+ concentration induced by ionomycin, a Ca2+ ionophore, and thapsigargin, which releases intracellular stores of that cation, inhibited NO-stimulated intracellular cGMP accumulation. Similarly, carbachol-induced elevation of the intracellular Ca2+ concentration inhibited NO-stimulated intracellular cGMP accumulation in HEK 293 cells. These data demonstrate that SGC behaves as a sensitive Ca2+ detector that may play a central role in coordinating the reciprocal regulation of Ca2+- and cGMP-dependent signaling mechanisms.


Subject(s)
Calcium/physiology , Guanylate Cyclase/metabolism , Nitric Oxide/physiology , Recombinant Proteins/metabolism , Animals , Binding, Competitive , Cell Extracts/antagonists & inhibitors , Cell Line , Cyclic GMP/antagonists & inhibitors , Enzyme Inhibitors/pharmacology , Guanine Nucleotides/physiology , Guanosine Triphosphate/metabolism , Guanylate Cyclase/antagonists & inhibitors , Guanylate Cyclase/biosynthesis , Guanylate Cyclase/genetics , Humans , Intracellular Fluid/metabolism , Kidney/cytology , Nitric Oxide/antagonists & inhibitors , Rats , Recombinant Proteins/antagonists & inhibitors , Recombinant Proteins/biosynthesis , Solubility
19.
Endocrinology ; 140(6): 2555-61, 1999 Jun.
Article in English | MEDLINE | ID: mdl-10342842

ABSTRACT

We have recently reported that members of the bcl-2 gene family are expressed and estradiol regulated in rabbit luteal cells during corpus luteum (CL) regression, and that estradiol and hCG are effective inhibitors of apoptosis in the rabbit CL in vivo and in vitro. As Bcl-2 and related proteins are known to regulate levels of reactive oxygen species or their intermediates in cells as one possible mechanism to control apoptosis, the present studies were designed to examine if oxidative stress plays a role in luteal cell apoptosis during CL regression in the rabbit. In the first set of experiments, healthy CL obtained from day 11 pseudopregnant rabbits were incubated in serum-free medium for 2 h in the absence or presence of superoxide dismutase (SOD; 1.5-150 U/ml), ascorbic acid (1-100 mM), N-acetyl-L-cysteine (25 and 50 mM), or catalase (10-1000 U/ml). Cells within CL incubated in medium alone exhibited extensive apoptosis (examined by analysis of extracted DNA using 3'-end labeling), and this onset of apoptosis was blocked in a dose-dependent fashion by treatment with SOD, ascorbic acid, N-acetyl-L-cysteine, or catalase. In the second set of experiments, expression of bax and bcl-x in CL after in vitro treatment without and with 100 U/ml SOD was examined. Although SOD treatment did not alter the levels of bcl-x messenger RNA (mRNA) over the 2-h incubation period, this antioxidant enzyme significantly reduced the levels of bax mRNA in incubated CL. In the final set of experiments, we observed that expression of mitochondrial- or manganese-containing SOD was significantly increased by treatment of isolated CL with 1 microg/ml hCG in vitro, whereas bax mRNA levels were significantly reduced under the same culture conditions. Collectively, these data indicate that the gonadotropin-mediated inhibition of apoptosis in rabbit luteal cells involves enhanced expression of the oxidative stress response gene, manganese-containing SOD, whose protein product may then function to protect luteal cells directly from the damaging effect of reactive oxygen species and/or indirectly by acutely down-regulating expression of Bax, a prooxidant member of the Bcl-2 protein family.


Subject(s)
Antioxidants/pharmacology , Apoptosis/drug effects , Chorionic Gonadotropin/pharmacology , Corpus Luteum/drug effects , Gene Expression Regulation , Proto-Oncogene Proteins/genetics , Superoxide Dismutase/physiology , Animals , Corpus Luteum/metabolism , Corpus Luteum/pathology , Female , Proto-Oncogene Proteins c-bcl-2/genetics , Pseudopregnancy/metabolism , Rabbits , Reactive Oxygen Species , bcl-2-Associated X Protein , bcl-X Protein
20.
FASEB J ; 13(8): 913-22, 1999 May.
Article in English | MEDLINE | ID: mdl-10224234

ABSTRACT

Bacteria that produce heat-stable enterotoxins (STs), a leading cause of secretory diarrhea, are a major cause of morbidity and mortality worldwide. ST stimulates guanylyl cyclase C (GCC) and accumulation of intracellular cyclic GMP ([cGMP]i), which opens the cystic fibrosis transmembrane conductance regulator (CFTR)-related chloride channel, triggering intestinal secretion. Although the signaling cascade mediating ST-induced diarrhea is well characterized, antisecretory therapy targeting this pathway has not been developed. 2-ChloroATP (2ClATP) and its cell-permeant precursor, 2-chloroadenosine (2ClAdo), disrupt ST-dependent signaling in intestinal cells. However, whether the ability to disrupt guanylyl cyclase signaling translates into effective antisecretory therapy remains untested. In this study, the efficacy of 2ClAdo to prevent ST-induced water secretion by human intestinal cells was examined. In Caco-2 human intestinal cells, ST increased [cGMP]i, induced a chloride current, and stimulated net basolateral-to-apical water secretion. This effect on chloride current and water secretion was mimicked by the cell-permeant analog of cGMP, 8-bromo-cGMP. Treatment of Caco-2 cells with 2ClAdo prevented ST-induced increases in [cGMP]i, chloride current and water secretion. Inhibition of the downstream consequences of ST-GCC interaction reflects proximal disruption of cGMP production because 8-bromo-cGMP stimulated chloride current and water secretion in 2ClAdo-treated cells. Thus, this study demonstrates that disruption of guanylyl cyclase signaling is an effective strategy for antisecretory therapy and provides the basis for developing mechanism-based treatments for enterotoxigenic diarrhea.


Subject(s)
Bacterial Toxins/toxicity , Diarrhea/drug therapy , Diarrhea/etiology , Enterotoxins/toxicity , Signal Transduction/drug effects , 2-Chloroadenosine/pharmacology , Biological Transport, Active/drug effects , Caco-2 Cells , Cell Differentiation , Chlorides/metabolism , Cystic Fibrosis Transmembrane Conductance Regulator/drug effects , Cystic Fibrosis Transmembrane Conductance Regulator/metabolism , Diarrhea/physiopathology , Enzyme Inhibitors/pharmacology , Escherichia coli Proteins , Guanylate Cyclase/antagonists & inhibitors , Guanylate Cyclase/genetics , Guanylate Cyclase/metabolism , Humans , Ion Transport/drug effects , Phosphorylation , Prodrugs/pharmacology , Signal Transduction/physiology , Water/metabolism
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