Cysteine protects freshly isolated cardiomyocytes against oxidative stress by stimulating glutathione peroxidase

Cysteine has been implicated in myocardial protection, although this is controversial and constrained by limited knowledge about the effects of cysteine at the cellular level. This study tested the hypothesis that a physiologically relevant dose of l-cysteine could be safely loaded into isolated cardiomyocytes leading to improved protection against oxidative stress. Freshly isolated adult rat ventricular cardiomyocytes were incubated for 2 h at 37°C with (cysteine incubated) or without (control) 0.5 mM cysteine prior to washing and suspension in fresh cysteine-free media. Cysteine incubated cells had higher intracellular cysteine levels compared to controls (9.6 ± 0.78 vs. 6.5 ± 0.65 nmol/mg protein, P < 0.02, n = 6 ± SE). Cell homeostasis indicators were similar in the two groups. Cysteine incubated cells had significantly higher glutathione peroxidase (GPx) activity (1.11 ± 0.23 vs. 0.54 ± 0.1 U/mg protein, P < 0.05, n = 5 ± SE) and significantly greater expression of GPx-1 (5.01 ± 0.48 vs. 3.01 ± 0.25 OD units/mm², P < 0.05, n = 4 ± SE) compared to controls. Upon exposure to H₂O₂, cysteine incubated cells generated fewer reactive oxygen species and took longer to show contractile changes and undergo hypercontracture. However, when cells were exposed to H₂O₂ in the presence of 0.05 mM of the GPx inhibitor mercaptosuccinic acid, this increased the control cells’ susceptibility to H₂O₂ and completely abolished the cysteine mediated protection. These results suggest a new role for cysteine in myocardial protection involving stimulation of glutathione peroxidase.     

Purification of xanthine dehydrogenase from rat liver: a rapid procedure with high enzyme yields

Xanthine dehydrogenase (EC 1.2.1.37) was purified approximately 1000-fold from liver homogenates of adult male Sprague-Dawley rats. Enzyme recovery was good (greater than 20% of the starting activity was obtained), and the homogeneously pure enzyme had a molecular mass of approximately 300,000 Da. The purified protein exhibited a specific activity of 2470 units/mg protein and spectral properties identical to those of the best preparations of this enzyme reported by other investigators. Routine preparations of this enzyme also possess higher dehydrogenase:oxidase ratios (typically between 5 and 6) than do other xanthine dehydrogenase preparations so far reported in the literature. Maximum dehydrogenase:oxidase ratios, greater than 10, could be obtained from this procedure if only peak dehydrogenase fractions from the chromatography columns were saved. The present small-scale purification method, which can be completed in 48-60 h, utilizes ammonium sulfate fractionation, Sephadex G-200 column chromatography, Blue Dextran-Sepharose column chromatography, and preparative gel electrophoresis.     

Rac1 Activation in Podocytes Induces Rapid Foot Process Effacement and Proteinuria

The kidney''s vital filtration function depends on the structural integrity of the glomerulus, the proximal portion of the nephron. Within the glomerulus, the architecturally complex podocyte forms the final cellular barrier to filtration. Injury to the podocyte results in a morphological change called foot process effacement, which is a ubiquitous feature of proteinuric diseases. The exact mechanism underlying foot process effacement is not known, but recently it has been proposed that this change might reflect activation of the Rac1 GTPase. To test this hypothesis, we generated a podocyte-specific, inducible transgenic mouse line that expressed constitutively active Rac1. When the Rac1 transgene was induced, we observed a rapid onset of proteinuria with focal foot process effacement. Using superresolution imaging, we verified that the induced transgene was expressed in damaged podocytes with altered foot process morphology. This work sheds new light on the complex balance of Rho GTPase signaling that is required for proper regulation of the podocyte cytoskeleton.     

Interference of seed dressing on groundnut nodulation

Aldrex T enhanced the mean number of groundnut nodules due to in vitro application at a higher than field concentration (6 g/kg groundnut kernels). Dressing at lower and agricultural applications (1.5 and 3, respectively) not only depressed but also considerably delayed nodule formation. This feature was demonstrated by the great losses in nodule number and weight as well. The latter picked up after 11 weeks to equate that of the controls. Interference of the fungicide in nodulation was highly significant at the sixth week of sowing. An insignificant increase in the number of root nodules was recorded at the tenth week of plant growth thus signifying recovery ofRhizobium from aldrex sensitivity. Dressing at 6 g/kg seeds did not affect the number and weight of nodules particularly after the fourth week of groundnut growth. Subsequently a highly significant increase in nodule weight was noticed.Nitrate and nitrite yield of nodules was favourably affected when kernels received twice the agricultural dose when compared with the non-dressed controls.     

Sensitivity of the root nodule rhizobium leguminosarum to the anti-fungal dressing aldrex T

Induction of inhibited growth halo, around filter paper discs or groundnut seeds dusted with aldrex T, diameter of which consistently increased with dressing application rate, was exhibited by the root nodule bacteriumRhizobium leguminosarum. Some cells sensitive to aldrex T gained recovery upon transference to fresh media denoting a bacteriostatic effect of the anti-fungal dressing.The viable bacterial cells steadily declined with fungicide concentrations between 100 and 500 p.p.m. An increase of 130% in viable colony count was recorded when the dose was increased to 700 p.p.m., and a relatively similar increase at 900 p.p.m., thus exhibiting a zone of inverted growth at 500 p.p.m. which was also confirmed by the optical density test. The latter was reliable after 24h incubation when the turbidity uniformly deminished with the fungicide dose to a maximal decline at 500 p.p.m. Aldrex T decreased the log and increased the stationary phases ofR. leguminosarum growth at all test doses.In vitro treatment ofR. leguminosarum with the fungicide exhibited some changes in bacterial morphology, cell lysis and shrinkage of cell membrane as revealed by the electron microscope.     

Depression comorbidity among patients with tuberculosis in a university teaching hospital outpatient clinic in Nigeria

Background Tuberculosis (TB) remains a leading infectious cause of morbidity and mortality throughout the world. Medication non-compliance has been recognised as one of the drawbacks in the successful management of this disease. Hence, different approaches for ensuring medication compliance have been adopted; these include the Directly Observed Therapy Short course (DOTS). TB is associated with psychiatric morbidity, particularly depressive disorder, and this has been recognised as a cause of poor compliance and a cause of increased morbidity and mortality from the disease. Despite this recognition, little attention is paid to the identification of depression among TB patients, particularly in the DOTS clinics that most of these patients attend. This study was designed to determine the prevalence of depression in patients with TB attending the DOTS outpatient clinic in a university teaching hospital in Nigeria, and to find out the factors that may be associated with this.Method All consenting TB patients attending the clinic completed a socio-demographic questionnaire and nine-item Patient Health Questionnaire (PHQ-9) designed to screen for depression, especially in outpatient and primary care settings.Results Sixty-five patients participated in the study of whom 41 (63.1%) were males. The mean age of the respondents was 35.1 ± 14.4 (range 15-70 years). Eighteen (27.7%) of the patients had depression, comprising 14 (21.5%) with mild depression and four (6.2%) with moderate depression. Socio-demographic factors (age groups, P=0.024; and financial status, P=0.02) and a clinical factor (persistent cough, P=0.04) were significantly associated with depression.Conclusion Measures to reduce depression among patients with TB should include effective symptom control, particularly of coughing, and measures to improve the financial status of this group of patients. Financial empowerment of patients may reduce depression in them, improve the compliance rate to anti-TB medication, and could furthermore bring an improvement to their quality of life.     

Understanding the Role of Accredited Drug Dispensing Outlets in Tanzania’s Health System

IntroductionPeople in many low-income countries access medicines from retail drug shops. In Tanzania, a public-private partnership launched in 2003 used an accreditation approach to improve access to quality medicines and pharmaceutical services in underserved areas. The government scaled up the accredited drug dispensing outlet (ADDO) program nationally, with over 9,000 shops now accredited. This study assessed the relationships between community members and their sources of health care and medicines, particularly antimicrobials, with a specific focus on the role ADDOs play in the health care system.MethodsUsing mixed methods, we collected data in four regions. We surveyed 1,185 households and audited 96 ADDOs and 84 public/nongovernmental health facilities using a list of 17 tracer drugs. To determine practices in health facilities, we interviewed 1,365 exiting patients. To assess dispensing practices, mystery shoppers visited 306 ADDOs presenting one of three scenarios (102 each) about a child’s respiratory symptoms.ConclusionADDOs are the principal source of medicines in Tanzania and an important part of a multi-faceted health care system. Poor prescribing in health facilities, poor dispensing at ADDOs, and inappropriate patient demand continue to contribute to inappropriate medicines use. Therefore, while accreditation has attempted to address the quality of pharmaceutical services in private sector drug outlets, efforts to improve access to and use of medicines in Tanzania need to target ADDOs, public/nongovernmental health facilities, and the public to be effective.     

Assembly of an SAP97-AKAP79-cAMP-dependent protein kinase scaffold at the type 1 PSD-95/DLG/ZO1 motif of the human beta(1)-adrenergic receptor generates a receptosome involved in receptor recycling and networking

Appropriate trafficking of the beta(1)-adrenergic receptor (beta(1)-AR) after agonist-promoted internalization is crucial for the resensitization of its signaling pathway. Efficient recycling of the beta(1)-AR required the binding of the protein kinase A anchoring protein-79 (AKAP79) to the carboxyl terminus of the beta(1)-AR (Gardner, L. A., Tavalin, S. A., Goehring, A., Scott, J. D., and Bahouth, S. W. (2006) J. Biol. Chem. 281, 33537-33553). In this study we show that AKAP79 forms a complex with the type 1 PDZ-binding sequence (ESKV) at the extreme carboxyl terminus of the beta(1)-AR, which is mediated by the membrane-associated guanylate kinase (MAGUK) protein SAP97. Thus, the PDZ and its associated SAP97-AKAP79 complex are involved in targeting the cyclic AMP-dependent protein kinase (PKA) to the beta(1)-AR. The PDZ and its scaffold were required for efficient recycling of the beta(1)-AR and for PKA-mediated phosphorylation of the beta(1)-AR at Ser(312). Overexpression of the catalytic subunit of PKA or mutagenesis of Ser(312) to the phosphoserine mimic aspartic acid both rescued the recycling of the trafficking-defective beta(1)-ARDelta PDZ mutant. Thus, trafficking signals transmitted from the PDZ-associated scaffold in the carboxyl terminus of the beta(1)-AR to Ser(312) in the 3rd intracellular loop (3rd IC) were paramount in setting the trafficking itinerary of the beta(1)-AR. The data presented here show that a novel beta(1)-adrenergic receptosome is organized at the beta(1)-AR PDZ to generate a scaffold essential for trafficking and networking of the beta(1)-AR.