ABSTRACT
Streptococcus mutans is a component of the dental plaque biofilm and a major causal agent of dental caries. Log-phase cells of the organism are known to induce an acid tolerance response (ATR) at sub-lethal pH values ( approximately 5.5) that enhances survival at lower pH values such as those encountered in caries lesions. In this study, we have employed a rod biofilm chemostat system to demonstrate that, while planktonic cells induced a strong ATR at pH 5.5, biofilm cells were inherently more acid resistant than such cells in spite of a negligible induction of an ATR. Since these results suggested that surface growth itself triggered an ATR in biofilm cells, we were interested in comparing the effects of a pH change from 7.5 to 5.5 on protein synthesis by the two cell types. For this, cells were pulse labeled with [(14)C]-amino acids following the pH change to pH 5.5, the proteins extracted and separated by two-dimensional (2D) electrophoresis followed by autoradiography and computer-assisted image analysis. A comparison between the cells incubated at pH 5.5 and the control biofilm cells revealed 23 novel proteins that were absent in the control cells, and 126 proteins with an altered relative rate of synthesis. While the number of changes in protein expression in the biofilm cells was within the same range as for planktonic cells, the magnitude of their change was significantly less in biofilm cells, supporting the observation that acidification of biofilm cells induced a negligible ATR. Mass spectrometry and computer-assisted protein sequence analysis revealed that ATR induction of the planktonic cells resulted in the downregulation of glycolytic enzymes presumably to limit cellular damage by the acidification of the external environment. On the other hand, the glycolytic enzymes in control biofilm cells were significantly less downregulated and key enzymes, such as lactate dehydrogenase were upregulated during pH 5.5 incubation, suggesting that the enhanced acid resistance of biofilm cells is associated with the maintenance of pH homeostasis by H+ extrusion via membrane ATPase and increased lactate efflux.
Subject(s)
Acids/pharmacology , Biofilms/drug effects , Streptococcus mutans/drug effects , Adaptation, Physiological , Biofilms/growth & development , Electrophoresis, Gel, Two-Dimensional , Heat-Shock Response , Hydrogen-Ion Concentration , Mouth/microbiology , Streptococcus mutans/physiologyABSTRACT
AIMS/HYPOTHESIS: Type I (insulin-dependent) diabetes mellitus is characterized by selective destruction of the insulin producing beta cells. Interleukin-1beta (IL-1beta) modulates the beta-cell function, protein synthesis, energy production and causes apoptosis. We have previously shown changes in the expression of 82 out of 1 815 protein spots detected by two dimensional gel electrophoresis in IL-1beta exposed diabetes prone Bio Breeding (BB-DP) rat islets of Langerhans in vitro. The aim of this study was to identify the proteins in these 82 spots by mass spectrometry and compare these changes with those seen in IL-1beta exposed Wistar Furth (WF) rat islets. METHODS: The 82 protein spots, that changed expression after IL-1beta exposure, were all re-identified on preparative gels of 200 000 neonatal WF rat islets, cut out and subjected to mass spectrometry for identification. RESULTS: Forty-five different proteins were identified from 51 spots and grouped according to function: (i) energy transduction and redox potentials; (ii) glycolytic and Krebs cycle enzymes; (iii) protein, DNA and RNA synthesis, chaperoning and protein folding; (iv) signal transduction, regulation, differentiation and apoptosis; (v) cellular defence; and (vi) other functions. Comparison of IL-1beta exposed BB-DP and WF islets showed common changes in 14 proteins and several proteins influencing similar pathways, suggesting that similar routes in the two strains lead to beta-cell destruction. CONCLUSION/INTERPRETATION: We demonstrate that proteome analysis is a powerful tool to identify proteins and pathways in BB-DP rat islets exposed to IL-1beta.
Subject(s)
Diabetes Mellitus, Type 1/genetics , Interleukin-1/pharmacology , Islets of Langerhans/physiology , Proteins/genetics , Proteome , Animals , Animals, Newborn , Cells, Cultured , Electrophoresis, Gel, Two-Dimensional , Enzymes/genetics , Islets of Langerhans/cytology , Islets of Langerhans/drug effects , Proteins/isolation & purification , Rats , Rats, Inbred BB , Rats, Inbred WFABSTRACT
Our study aimed to evaluate the influence of thrombolytic therapy on some left ventricle (LV) function parameters in patients with acute myocardial infarction. The study was performed on 44 pts admitted to hospital due to acute myocardial infarction. The patients were divided into two groups: I group--30 pts (26 male, 4 female) at average age 57 +/- 10 who were treated with tissue plasminogen activator (t-PA) routinely and II group--14 pts (9 male, 5 female) at average age 62 +/- 10 in whom thrombolytic therapy was contraindicated for various reasons. Transthoracic echocardiography was performed just before treatment (0), 3.5 hours after the onset of drug administration (2 hours after the end of t-PA injection) (1) and on the 10th day of hospitalization (2). Control group consisted of 16 clinically healthy individuals (12 male, 4 female) at average age 54 +/- 9. The following parameters were evaluated: DT-E--wave of early diastolic transmitral flow deceleration time, IVRT--isovolumic relaxation time, E/A--early/atrial peak flow velocity ratio of transmitral flow, LATEF%--left atrial total emptying fraction, EF--left ventricle ejection fraction. In patients with acute myocardial infarction shortening of DT, prolongation of IVRT, lower E/A ratio and decrease of LATEF% compared to controls were observed. In group I EF was less than in clinically healthy individuals. E/A ratio was higher in pts from group I than from group II. In patients treated with t-PA 2 hours after treatment as well as on the 10th day significant prolongation of DT, shortening of IVRT and increase of LATEF% were observed. These changes were accompanied by the increase of EF. In patients with acute myocardial infarction not treated with t-PA significant increase in E/A ratio and EF on 10th day were observed. On the basis of the results were conclude: In patients with acute myocardial infarction LV diastolic function and with unproper relaxation as well as unproper compliance of LV myocardium is present. In patients with thrombolytic therapy LV filling pattern improves just two hours after t-PA administration (DT prolongation, IVRT shortening, LATEF% increase). Such tendency remains on the 10th day after treatment. In patients without thrombolytic therapy slight improvement occurs no sooner than on the 10th day of the MI.
