Evolving concepts in biofilm infections

Several pathogens associated with chronic infections, including Pseudomonas aeruginosa in cystic fibrosis pneumonia, Haemophilus influenzae and Streptococcus pneumoniae in chronic otitis media, Staphylococcus aureus in chronic rhinosinusitis and enteropathogenic Escherichia coli in recurrent urinary tract infections, are linked to biofilm formation. Biofilms are usually defined as surface-associated microbial communities, surrounded by an extracellular polymeric substance (EPS) matrix. Biofilm formation has been demonstrated for numerous pathogens and is clearly an important microbial survival strategy. However, outside of dental plaques, fewer reports have investigated biofilm development in clinical samples. Typically biofilms are found in chronic diseases that resist host immune responses and antibiotic treatment and these characteristics are often cited for the ability of bacteria to persist in vivo . This review examines some recent attempts to examine the biofilm phenotype in vivo and discusses the challenges and implications for defining a biofilm phenotype.     

Evolving concepts of human state dissociation

The concept of state dissociation in humans was made possible only by applying information obtained from basic science animal research studies to the human condition--without which these often dramatic, and treatable conditions would have remained in the mystical, supra-natural, or psychiatric arenas, without appropriate or effective treatment options. Sleep or wakefulness occurring asynchronously in bits and pieces of the brain is a most useful concept. From our standpoint, the basic science work in the function and mechanism of sleep is pertinent, not only adding to our knowledge in these important areas for the sake of knowledge, but also in providing clinicians with important information that is of immense clinical importance. The payoff of such research has been great, and demands that it should be ongoing. The field of sleep research and sleep medicine is in a unique position to foster close interactions between basic scientists and clinicians, the result being basic science answers to clinical questions, and unanswered clinical questions guiding the direction of and reinforcing the basic science research. The clinical conditions discussed above underscore the value of close cooperation among those working at all levels: molecular, cellular, multi-cellular, and clinical. Continued study of state dissociation by both basic scientists and clinicians will undoubtedly identify and explain even more of these fascinating conditions, with important therapeutic implications. The reciprocal benefits of close collaboration between basic scientists and clinicians will continue to be realized.     

Evolving concepts of thyroid hormone action.

The past 25 years have witnessed dramatic changes in our concepts of thyroid hormone action. Progress in this area was made possible by the recognition of the central role of triiodothyronine in mediating thyroid hormone action and the recognition of specific nuclear receptors in target tissues as demonstrated by displacement studies. The cloning of the receptors and receptor variants has enabled investigators to undertake detailed analyses of the biochemical events which underlie the physiological and pathological action of thyroid hormone.     

Evolving concepts in cancer therapy through targeting sphingolipid metabolism

Traditional methods of cancer treatment are limited in their efficacy due to both inherent and acquired factors. Many different studies have shown that the generation of ceramide in response to cytotoxic therapy is generally an important step leading to cell death. Cancer cells employ different methods to both limit ceramide generation and to remove ceramide in order to become resistant to treatment. Furthermore, sphingosine kinase activity, which phosphorylates sphingosine the product of ceramide hydrolysis, has been linked to multidrug resistance, and can act as a strong survival factor. This review will examine several of the most frequently used cancer therapies and their effect on both ceramide generation and the mechanisms employed to remove it. The development and use of inhibitors of sphingosine kinase will be focused upon as an example of how targeting sphingolipid metabolism may provide an effective means to improve treatment response rates and reduce associated treatment toxicity. This article is part of a Special Issue entitled Tools to study lipid functions.     

Evolving concepts in plant glycolysis: two centuries of progress

Glycolysis, the process responsible for the conversion of monosaccharides to pyruvic acid, is a ubiquitous feature of cellular metabolism and was the first major biochemical pathway to be well characterized. Although the majority of glycolytic enzymes are common to all organisms, the past quarter of a century has revealed that glycolysis in higher plants possesses numerous distinctive features. Research in the nineteenth century established convincingly that plants carry out alcoholic fermentation under anaerobic conditions. In 1878, Wilhelm Pfeffer asserted that a non-oxygen-requiring ‘intramolecular respiration’ was involved in the aerobic respiration of plants. Between 1900 and 1950 it was demonstrated that plants metabolize sugar and starch by a glycolytic pathway broadly similar to that of yeasts and muscle tissue. In 1948, the first purification and characterization of a plant glycolytic enzyme, aldolase, was published by Paul Stumpf. By 1960 the presence of each of the 10 enzymes of glycolysis, presumed at the time to be located in the cytosol, had been confirmed in higher plants. Shortly after 1960 it was shown that the mechanism of glycolytic regulation in plants had features in common with that of animals and yeasts, especially as regards the important role played by the enzyme phosphofructokinase; but important regulatory properties peculiar to plants were soon demonstrated. In the last 30 years, higher-plant glycolysis has been found to exhibit a number of additional characteristics peculiar to plant systems. One conspicuous feature of plant glycolysis, discovered in the 1970s, is the presence of a complete or nearly complete sequence of glycolytic enzymes in plastids, distinct and spatially separated from the glycolytic enzymes located in the cytosol. Plastidic and cytosolic isoenzymes of glycolysis have been shown to differ in their kinetic and regulatory properties, suggesting that the two pathways are independently regulated. Since about 1980 it has become increasingly clear that the cytosolic glycolysis of plants may make use of several enzymes other than the conventional ones found in yeasts, muscle tissue and plant plastids: these enzymes include a pyrophosphate-dependent phosphofructokinase, a non-reversible and nonphosphorylating glyceraldehyde-3-phosphate dehydrogenase, a phosphoenolpyruvate phosphatase (vacuolar location) and a three-enzyme sequence able to produce pyruvate from phosphoenolpyruvate avoiding the pyruvate-kinase step. These non-conventional enzymes may catalyze glycolysis in the plant cytosol especially under conditions of metabolic stress. Experiments on transgenic plants possessing significantly elevated or reduced (reduced to virtually nil in some cases) levels of glycolytic enzymes are currently playing an important part in improving our understanding of the regulation of plant glycolysis; such experiments illustrate an impressive degree of flexibility in the pathway's operation. Plant cells are able to make use of enzymes bypassing or substituting for several of the conventional enzymic steps in the glycolytic pathway; the extent and conditions under which these bypasses operate are the subject of current research. The duplication of the glycolytic pathway in plants and the flexible nature of the pathway have possibly evolved in relation to the crucial biosynthetic role played by plant glycolysis beyond its function in energy generation; both functions must proceed if a plant is to survive under varying and often stressful environmental or nutritional conditions.     

Evolving concepts on the pathogenic mechanisms of aniridia related keratopathy

Heterozygosity for PAX6 deficiency (PAX6+/-) results in aniridia. Corneal changes in aniridia-related keratopathy (ARK) include corneal vascular pannus formation, conjunctival invasion of the corneal surface, corneal epithelial erosions and epithelial abnormalities, which eventually result in corneal opacity and contribute to visual loss. Corneal changes in aniridia have been attributed to congenital deficiency of corneal limbal stem cells. The aim of this paper is to review the potential mechanisms that may underlie the pathogenesis of aniridia related keratopathy. Current evidence, based on clinical observations and an animal model of aniridia suggest that the proliferative potential of the corneal limbal stem cells may not primarily be impaired. The corneal changes in aniridia may be related to an abnormality within the limbal stem cell niche. The mechanisms underlying progressive corneal pathology in aniridia appear multi-factorial and include: (1) abnormal corneal healing responses secondary to anomalous extracellular matrix metabolism; (2) abnormal corneal epithelial differentiation leading to fragility of epithelial cells; (3) reduction in cell adhesion molecules in the PAX6 heterozygous state, rendering the cells susceptible to natural shearing forces; and (4) conjunctival and corneal changes leading to the presence of cells derived from conjunctiva on the corneal surface.     

Molecular heterogeneity of tumor endothelium

Tumors depend on a vascular supply for their growth. Tumor blood vessels, which are derived from normal tissue vasculature, display a markedly abnormal phenotype. Tumor endothelial properties are highly varied in space and time. An important goal is to delineate the range of phenotypes in tumor endothelium and to identify tumor endothelial-specific molecular signatures. This information should lead to a more complete understanding of the mechanisms of tumor growth, the discovery of new therapeutic targets, and the development of biomarkers for diagnosis and surveillance. The goals of this review are to outline recent advances in dissecting tumor endothelial-cell-specific gene expression, to address mechanisms of phenotypic heterogeneity in tumor vascular beds, and to discuss the therapeutic implications of these findings.     

Evolving concepts of the epidemiology, diagnosis, and therapy of Mycobacterium tuberculosis infection

Tuberculosis in the United States is evolving in nearly all respects--epidemiology, diagnosis, treatment, and prophylaxis. Today a relatively larger segment of the population has predisposing factors to infection with tuberculosis. There is a greater percentage of people who are elderly, who have immigrated from countries endemic for tuberculosis, or who are immunosuppressed due to medications necessary for other conditions, because of malignancies, or because of infection with HIV. Skin test classifications have been revised to give different meanings to different-sized areas of induration at the injection site for defined populations. More sensitive, more specific, and faster diagnostic laboratory tests for tuberculosis are being developed. Short-course chemotherapy of from six to nine months is now accepted as standard treatment, regardless of exactly which of the proven regimens of antibiotics or accepted lengths of therapy is used. Patient compliance is improved with the shorter courses both for treatment and for prophylaxis. Better compliance with therapy results in better treatment outcomes of infections with Mycobacterium tuberculosis.     

Intratumor heterogeneity in evolutionary models of tumor progression

With rare exceptions, human tumors arise from single cells that have accumulated the necessary number and types of heritable alterations. Each such cell leads to dysregulated growth and eventually the formation of a tumor. Despite their monoclonal origin, at the time of diagnosis most tumors show a striking amount of intratumor heterogeneity in all measurable phenotypes; such heterogeneity has implications for diagnosis, treatment efficacy, and the identification of drug targets. An understanding of the extent and evolution of intratumor heterogeneity is therefore of direct clinical importance. In this article, we investigate the evolutionary dynamics of heterogeneity arising during exponential expansion of a tumor cell population, in which heritable alterations confer random fitness changes to cells. We obtain analytical estimates for the extent of heterogeneity and quantify the effects of system parameters on this tumor trait. Our work contributes to a mathematical understanding of intratumor heterogeneity and is also applicable to organisms like bacteria, agricultural pests, and other microbes.     

Estimation of tumor heterogeneity using CGH array data

Background  

Array-based comparative genomic hybridization (CGH) is a commonly-used approach to detect DNA copy number variation in whole genome-wide screens. Several statistical methods have been proposed to define genomic segments with different copy numbers in cancer tumors. However, most tumors are heterogeneous and show variation in DNA copy numbers across tumor cells. The challenge is to reveal the copy number profiles of the subpopulations in a tumor and to estimate the percentage of each subpopulation.     

Allele-specific copy number analysis of tumor samples with aneuploidy and tumor heterogeneity

We describe a bioinformatic tool, Tumor Aberration Prediction Suite (TAPS), for the identification of allele-specific copy numbers in tumor samples using data from Affymetrix SNP arrays. It includes detailed visualization of genomic segment characteristics and iterative pattern recognition for copy number identification, and does not require patient-matched normal samples. TAPS can be used to identify chromosomal aberrations with high sensitivity even when the proportion of tumor cells is as low as 30%. Analysis of cancer samples indicates that TAPS is well suited to investigate samples with aneuploidy and tumor heterogeneity, which is commonly found in many types of solid tumors.     

Allele-specific copy number analysis of tumor samples with aneuploidy and tumor heterogeneity

We describe a bioinformatic tool, Tumor Aberration Prediction Suite (TAPS), for the identification of allele-specific copy numbers in tumor samples using data from Affymetrix SNP arrays. It includes detailed visualization of genomic segment characteristics and iterative pattern recognition for copy number identification, and does not require patient-matched normal samples. TAPS can be used to identify chromosomal aberrations with high sensitivity even when the proportion of tumor cells is as low as 30%. Analysis of cancer samples indicates that TAPS is well suited to investigate samples with aneuploidy and tumor heterogeneity, which is commonly found in many types of solid tumors.     

Mammary tumor heterogeneity in wt-ErbB-2 transgenic mice

Phenotypic and biological heterogeneity was studied in a single transgenic mouse model to determine the level of biological variance. We analyzed 1,258 tumors from 417 MMTV-wt-ErbB-2 transgenic mice, subdivided by casein or soy-based dietary randomization and hormonal treatment. Variance in tumor histologic features, growth pattern, invasion, metastases, and multi-focality were detected in untreated and treated mice. Ninety-three percent (1,174/1,258) of tumors had the solid growth pattern widely reported in this model. However, among the solid tumors, a spectrum of growth patterns, from well-circumscribed tumors with a pseudocapsule to locally invasive or highly aggressive, metastatic subtype, was observed. Of the non-solid tumors, glandular features were prominent in 84 (7%). Adenocarcinomas included papillary, acinar/glandular, and adenosquamous subtypes. Adenosquamous tumors were exclusively observed in the group of mice treated on a short-term basis with estrogen. In contrast to the reported literature for this transgenic mouse model, mammary tumors were multifocal in the majority of cases (303 of 417 mice, or 73%). Results of this extensive study of a single transgenic model of mammary tumorigenesis indicate phenotypic and biological heterogeneity not previously associated with this transgenic mouse. These data support a complex, multistep process of carcinogenesis and clonal evolution, with biological and phenotypic variance similar to that observed in human mammary cancer development.     

Evolving concepts of the intrarenal renin-angiotensin system in health and disease: contributions of molecular biology.

Previous physiological and biochemical studies suggest the existence of an endogenous renin-angiotensin system (RAS) in the kidney. However, these data cannot exclude the contribution of the circulating RAS. Proof of the local synthesis of RAS components in the kidney has been obtained recently through the use of molecular biological techniques. Using Northern blot analysis, we have demonstrated the intrarenal expression of renin, angiotensinogen, and angiotensin-converting enzyme messenger RNAs. Employing in situ hybridization histochemistry, we have localized the intrarenal tissue sites of renin and angiotensinogen messenger RNA synthesis. Renin gene expression was found in cells of the juxtaglomerular apparatus. Angiotensinogen mRNA was primarily produced in the proximal convoluted tubule with lesser amounts in glomerular tufts and vasculature. These findings led us to hypothesize that the proximal tubule is a major site of renal Ang II synthesis and that locally synthesized Ang II might directly modulate tubular function. Both genes are subject to feedback regulation. Our studies showed that Ang II exerted a stimulatory effect on angiotensinogen but a negative feedback effect on renin gene expression. Dietary NaCl restriction stimulated the expression of both genes, although the onset of renin gene activation required more prolonged sodium chloride restriction. Furthermore, our data indicated that the sodium cation, irrespective of the anion, was primarily important in regulating renal angiotensinogen mRNA levels. Our studies also showed altered intrarenal renin or angiotensinogen expressions in pathophysiological states, e.g. in experimental heart failure and the spontaneously hypertensive rat. Taken together, these data support the existence of a intrarenal RAS and suggest its potential roles in the regulation of renal function in health and disease.     

Proteomic strategies to reveal tumor heterogeneity among urothelial papillomas

Proteomics and immunohistochemistry were used to reveal tumor heterogeneity among urothelial papillomas (UPs) with the long term goal of predicting their biological potential in terms of outcome. First, we identified proteins that were deregulated in invasive fresh lesions as compared with normal urothelium, and thereafter we immunostained UPs with a panel of antibodies against some of the markers. Twenty-two major proteins showing variations of 2-fold or more in at least one-third of the invasive lesions were selected. Specific antibodies against several of the proteins were obtained, but only a few reacted positively in immunostaining. A panel consisting of antibodies against keratinocytes (CKs) 5, 13, 18, and 20 and markers of squamous metaplasia (CKs 7, 8, and 14) was used to probe normal urothelium and 30 UPs collected during a period of five years. Four UPs showed a normal phenotype, whereas the rest could be grouped in five major types that shared aberrant staining with the CK20 antibody. Type 1 heterogeneity (n = 4) showed preferred staining of the umbrella cells with the CK8 antibody. Type 2 (n = 11) was typified by the staining of the basal and intermediate layers with the CK20 antibody. Type 3 (n = 7) was characterized by the predominant staining of the basal cell layer with the CK5 antibody. Type 4 (n = 1) showed areas of CK7 negative cells, whereas type 5 (n = 3) showed loss of staining of the basal cells with the CK20. 29% of the patients experienced recurrences, but none progressed to invasive disease. Patients harboring phenotypic alterations in the basal cell compartment (types 3 and 5) showed the highest number of recurrences (4/7 and 2/3, respectively), and all type 3 lesions progressed to a higher degree of dedifferentiation. Even though a long term prospective study involving a larger sample size is required to assess the biological potential of these lesions, we believe that this approach will prove instrumental for revealing early phenotypic changes in different types of cancer.     

Potential and limitations of current concepts regarding the response of clonal plants to environmental heterogeneity

Plant ecologists have spent considerable effort investigating the physiological mechanisms and ecological consequences of clonal growth in plants. One line of research is concerned with the response of clonal plants to environmental heterogeneity. Several concepts and hypotheses have been formulated so far, suggesting that intra-clonal resource translocation, morphological plasticity on different organizational levels (e.g. leaves, ramets, fragments), and other features of clonal plants may represent potentially adaptive traits enabling stoloniferous and rhizomatous species to cope better with habitat patchiness. Although each of these concepts contributes substantially to our understanding of the ecology of clonal species, it is difficult to combine them into a consistent theoretical framework. This apparent lack of conceptual coherence seems partly be caused by an uncritical use of the term habitat heterogeneity. Researchers have not always acknowledged the fact that heterogeneity may refer to a number of fundamentally different aspects of environmental variability (i.e. scale, contrast, predictability, temporal vs. spatial heterogeneity), and that each of these aspects may, on one hand, allow for the evolution of specific plant responses to heterogeneity and, on the other, severely constrain the viability of potentially adaptive traits. Since adaptive responses are operational only in a narrow range of conditions (delimited by external environmental conditions and constraints internal to plants) it seems imperative to clearly define the context and the limits within which concepts regarding clonal plants' responses to heterogeneity are valid. In this paper an attempt is made to review a number of these concepts and to try and identify the necessary conditions for them to be operational. Special attention is paid (1) to different aspects of environmental heterogeneity and how they may affect clonal plants, and (2) to possible constraints (e.g. sectoriality, perception of environmental signals, morphological plasticity) on plant responses to patchiness.     

Microvascular endothelial cell heterogeneity: general concepts and pharmacological consequences for anti-angiogenic therapy of cancer

Microvascular endothelial cells display a large degree of heterogeneity in function depending on their location in the vascular tree. The existence of organ-specific, microvascular-bed-specific, and even intravascular variations in endothelial cell gene expression emphasizes their high cell-to-cell variability, which is furthermore extremely adaptable to altering conditions. The ability of microvascular endothelial cells to respond dynamically to pathology-related microenvironmental changes is particularly apparent in tumor-growth-associated angiogenesis. An understanding of how they behave, how their behavior varies between and within tumors, and how their behavior is related to responsiveness to drugs is critical for the development of effective anti-angiogenic treatment strategies. In this review, we introduce some general issues concerning organ-imprinted microvascular heterogeneity and highlight the importance of studying microvascular endothelial cell behavior in an in vivo context. This is followed by an overview of state-of-the-art knowledge regarding the nature of the variation in microenvironmental conditions in pre-clinical and clinical tumors and their consequences for tumor endothelial behavior. We provide recent insights into the in vivo molecular activation status of the endothelium and, finally, outline our current understanding of the way that anti-angiogenic drugs affect tumor endothelial cells in relation to their anti-tumor effects.