With the advent of multislice CT more than a decade ago, multislice CT angiography has demonstrated a huge potential in the less invasive imaging of cardiovascular disease, especially in the diagnosis of coronary artery disease. The diagnostic accuracy of multislice CT angiography has been significantly augmented with the rapid technical developments ranging from the initial 4-slice, to the current 64-slice and 256 and 320-slice CT scanners. This is mainly demonstrated by the improved spatial and temporal resolution when compared to the earlier type of CT scanners. Traditionally, multislice CT angiography is acquired with retrospective ECG-gating with acquisition of volume data at the expense of increased radiation dose, since data is acquired at the entire cardiac cycle, although not all of them are used for postprocessing or reconstructions. Recently, there is an increasing trend of utilising prospective ECG-gating in cardiac imaging with latest multislice CT scanners (64 or more slices) with significant reduction of radiation dose when compared to retrospective ECG-gating method. However, there is some debate as to the diagnostic value of prospective ECG-gating in the diagnosis of coronary artery disease, despite its attractive ability to reduce radiation dose. This article will review the performance of retrospective ECG-gating in the diagnostic value of coronary artery disease, highlight the potential applications of prospective ECG-gating, and explore the future directions of multislice CT angiography in cardiac imaging. Coronary artery disease (CAD) is the leading cause of death in western countries. The standard of reference for diagnosis of CAD is still invasive coronary angiography, with the advantage of high spatial resolution and temporal resolution. Despite its cost, inconvenience to patients, and a small but distinct procedure-related morbidity (1.5%) and mortality (0.2%) rate, more than 1 million invasive diagnostic coronary angiography procedures are performed annually in the United States alone. Similarly, CAD is the single most important cause of death in Australia and New Zealand. Every year, billions of dollars have been spent in the treatment of coronary artery disease . Each year cardiovascular disease resulted in 81,926 coronary angiography examinations in Australia 2001-02 . Given the invasiveness of coronary angiography and potential danger of having a small risk of serious complications , a non-invasive technique for imaging of the coronary artery disease is highly desirable.Imaging of the heart and coronary artery branches has always been technically challenging due to the heart’s continuous movement. Over the last decade, great strides have been made in the field of cardiac imaging as non-invasive coronary imaging modalities have undergone rapid developments -4. InitiMultislice CT represents technical evolution in cardiac imaging when 4-slice CT scanner was first introduced into the clinical practice in 1998 . DiagnosRecently, prospective ECG-gating was introduced for cardiac MSCT angiography, and this imaging protocol is increasingly being reported in the literature, despite sufficient evidence still needed to verify its diagnostic value. This paper will review the diagnostic accuracy of retrospective gating CT angiography; the potential applications of prospective gating cardiac CT angiography, and highlight some future directions of MSCT in cardiac imaging.The feasibility of cardiac MSCT was initially demonstrated with 4-slice CT using retrospective ECG-gated technique. Volumetric CT data is acquired throughout the entire cardiac cycle during simultaneous recording of the ECG signal. Subsequently, data from specific periods of the cardiac cycle (most commonly at late diastolic phase) is reconstructed by retrospective referencing to the ECG signal with the aim of generating images with the least motion artefacts. Over the last decade a great deal of interest has been focused on imaging and diagnosis of CAD with MSCT due to its less invasive nature and fast scanning technique with extended z-axis coverage when compared to single slice CT. Earlier studies with 4-slice CT showed moderate diagnostic accuracy with pooled sensitivity and specificity of 78% and 93%, respectively . HoweverShorter examination times are possible with further improved diagnostic accuracy with 64-slice CT owing to improved spatial and temporal resolution compared with 16-slice CT. Acquisition of isotropic volume data are made available with 64-slice CT, thus detection of main and side coronary artery branches is improved when compared to earlier types of MSCT scanners. Several meta-analyses of 64-slice CT studies reported sensitivities of 93% and specificities of 96% (in 6 studies) , sensitiet al. [et al. in their recent study concluded that high diagnostic accuracy  was achieved even at high heart rates [In 2006, the first dual source multislice CT scanner was introduced . With thet al.  in theiret al. -22. RixeWhile satisfactory results have been achieved with the retrospective ECG-gating through continuous exposure in a low-helical pitch (0.2-0.4) resulting in multiple overlapping regions of X-ray exposure, the downside is the relatively high effective radiation doses. The radiation dose associated with retrospective ECG-gating is gradually increased with the increased number of detector rows and reduction of detector size. Thus, 4-slice CT scanners have lower dose than 16-slice scanners. Similarly 16-slice scanners have lower doses than 64-slice, and subsequently doses from 64-slice will be lower than those acquired with 256 and 320 slice scanners. Therefore, various strategies have been taken to reduce the radiation dose while using MSCT angiography in cardiac imaging, and prospective ECG-gating is by far the most effective and significant technique to reduce radiation dose.Prospective ECG-gating utilises the same technique as that used in electron-beam CT which is defined as the step and shoot method. It is mainly used for quantification of calcium burden, but recently it is increasingly used for CT coronary angiography examinations. The scan is performed in a non-helical way with acquisition of a series of axial images instead of volumetric data. Unlike retrospective gating, prospective ECG-gating or prospective triggering allows for acquisition of data by selectively turning the x-ray tube on only in the selected phase, triggered by the ECG signal, and turning off during the rest of R-R cycle. This is also referred to as sequential or step-and-shoot acquisition with prospective triggering and the effective pitch is 1.0. The main advantage of this scanning protocol is the lower radiation dose as X-ray exposure only takes place during the selected cardiac phase rather than throughout the entire cardiac cycle. Therefore, a significant reduction of radiation dose can be expected from prospective ECG-gating, which is the most attractive side of this scanning protocol compared to retrospective ECG-gating.Recent technical developments of MSCT imaging technique allows for prospective ECG-gating to be performed in a single heart beat with helical scan , 23, 24.et al. [et al. [Use of prospective ECG-gating with 64-slice or dual source CT has been reported to reduce the effective radiation dose by up to 90% when compared to retrospective ECG-gating technique -37. In 2et al.  first de [et al.  in their [et al. , 28. It  [et al. , 39, altet al. in their prospective study compared a group of patients who underwent prospectively gated cardiac CT with another matched group of patients who underwent retrospectively gated cardiac CT. Similar image quality of coronary segments was scored for both groups, but 77% radiation dose reduction was achieved in the prospectively gated group [et al. and Earls et al. who both showed the similar image quality between prospective and retrospective gated imaging, but with dose reduction of 79% and 83% achieved in the prospective groups [et al. reported their experience of using prospective gating in the largest clinical group including more than 2000 cases [Studies comparing prospective and retrospective gating further confirmed the significant reduction of radiation dose resulting from the former imaging protocol. Shuman ed group . This wae groups , 32. Moset al. studied the image quality and radiation dose with dual source CT prospective gating by using different protocols [et al. in their recent study consisting of 149 patients compared prospective gating and retrospective gating protocols with different kV groups [Radiation dose can be further reduced with reduction of the tube voltage (kVp), in addition to the tube current modulation which is available in most of the 64-slice scanners. Researchers investigated the prospective gating with different kVp, and its effect on radiation dose, and results are promising. Stolamann rotocols . Their rV groups . Their r2, lower image quality occurs as reported by recent studies [It is important to note that while prospective gating leads to a significant reduction in effective radiation dose and provides equivalent or improved image quality relative to retrospective gated images, studies reported in the literature highlight some important limitations to the current 64-slice CT . The main limitation lies in the fact that image quality is dependent on the heart rate, heart rate variation and body mass index (BMI). Maximum heart rate threshold is between 63-75 bmp for prospective gated imaging. When heart rate is greater than 70 bmp, heart rate variation greater than 10 bmp, or BMI greater than 30 kg/m studies , 36. Allet al. reported the superiority of 256 prospective gating CT in their initial experience with high image quality, low dose, independent of higher heart rates and higher BMI [et al. [Further technical developments in MSCT technique overcome the abovementioned limitations with the emergence of new generation of MSCT techniques such as 256- and 320-slice CT scanners. Longer z-axis coverage available with 256 and 320-slice scanners ranging from 12.8 cm to 16 cm in one gantry rotation permits full cardiac coverage in one gantry rotation with prospective gating, thus, eliminating the restrictions and limitations associated with 64-slice scanners , 39-41. gher BMI , 24. Theet al. presented the first report demonstrating the diagnostic performance of low-dose prospective gating CT for the diagnosis of CAD [et al. also reported the high diagnostic accuracy of prospective gating for diagnosis of CAD with low radiation dose, even in the presence of heavy calcification, despite the fact that increased rate of non-diagnostic segments was observed due to heavy calcification [Despite the promising aspect of significant reduction of radiation dose with prospective gating, there is lack of sufficient evidence to show the diagnostic value or performance of prospective gating in the detection of coronary artery disease. Most of the studies currently available in the literature addressed the image quality and reduction of radiation dose when comparing prospective with retrospective gated protocols. A direct comparison between prospective gating and conventional coronary angiography is scarce and data is limited for the diagnosis of coronary artery stenosis. Scheffel s of CAD . Diagnosfication . Furtheret al. noticed that 62% of patients have stair-step artefacts in prospective gated coronary CT angiography of coronary arteries [Apparently, there are two important limitations of prospective gating performed with 64-slice CT: prospective gating is limited to heart rates lower than 75 bpm due to short z-axis coverage. The z-axis coverage during image acquisition with 64-slice CT is only 4 cm, thus three to five dataset are required subsequently to image the entire heart. Stair-step artefacts due to misalignment of two adjacent datasets may happen if the position of the patient on the CT table changes during table travel, or heart rate is irregular or heart rate variation is great during the scan. Husmann arteries . The staet al. in their initial experience demonstrated that 320-slice CT allows for simultaneous perfusion imaging for the entire myocardium [The first limitation has been addressed with the use of 256 or 320 slice CT as shown by some studies mentioned above , 23, 24.ocardium , 45. AppIt seems that the current direction of using MSCT in cardiac imaging is moving from previous retrospective gating to prospective gating, and this is demonstrated by the increasing reports available in the literature over the last few years. The main driving force for this trend is the reduction of radiation dose, which is the most attractive aspect of prospective gating technique. Certainly this is only made available due to the technical developments of multislice CT technique, especially with increased temporal resolution. In order to acquire diagnostic quality images, prospective gating CT scans must be performed with use of 64- or more slice CT scanners. While radiation dose is significantly lower than that acquired with retrospective gating technique, the evidence of prospective gating for diagnosis of coronary artery disease is scarce at this stage. Thus, it is too early to draw conclusions that prospective gating can be used as a reliable alternative to conventional coronary angiography for the diagnosis of coronary artery disease before adequate research evidence is available.With more research findings available in the literature in the near future, it is expected that multislice CT will be used as a first line technique in cardiac imaging, possibly with prospective gating replacing the traditional retrospective gating technique. There is no doubt that multislice CT has entered a new era in cardiac imaging with the advent of 64-, 256- and 320-slice scanners, and its applications in clinical practice will benefit more patients suspected of coronary artery disease. While reduction of radiation dose is important, the most important aspect of cardiac CT is the image quality required for diagnostic purposes. Thus, both of these two factors need to be taken into account when choosing prospective gating CT coronary angiography in the diagnosis of coronary artery disease . The folWith use of 64-slice or dual source CT, in patients suspected of CAD for whom only the cardiac anatomy or presence or absence of CAD is the main concern, prospective gating is suggested;With use of 64-slice or dual source CT in patients suspected of CAD, if cardiac functional information will make a meaningful or significant contribution to the CT assessment or clinical treatment, use of retrospective gating is suggested with additional dose being justified;With use of 256-or 320-slice CT in cardiac imaging, prospective gating is recommended since it allows acquisition of dataset in one gantry rotation, thus providing both anatomical assessment and physiological evaluation;With use of prospective gated protocol, 100 kVp is recommended in patients with BMI less than 30 for further reduction of the effective radiation dose;Narrowing the phase window width in prospective gated protocol is recommended to reduce patient radiation dose in a single heartbeat CT angiography (256- or 320-slice CT).


In India, the Revised National Tuberculosis Control Programme (RNTCP) has adopted the strategy of examining two specimens during follow-up culture examinations to monitor the treatment response of multi-drug resistant tuberculosis (MDR-TB) patients.To determine the incremental yield of the second sputum specimen during follow-up culture examinations among patients with MDR-TB and the effect on case management on changing from two to one specimen follow-up strategy.A cross sectional record review of MDR-TB patients registered during 2008–09 under RNTCP was undertaken in three MDR-TB treatment sites of India.Of 1721 pairs of follow-up sputum culture examinations done among 220 MDR-TB patients, 451(26%) were positive with either of the two specimens; 29(1.7%) were culture positive only on the second specimen indicating the incremental yield. To detect one additional culture positive result on the second specimen, 59 specimens needed to be processed. If we had examined only one specimen, we would have missed 29 culture-positive results. By current RNTCP guidelines, however, a single specimen policy would have altered case management in only 3(0.2%) instances, where patients would have missed a one month extension of the intensive phase of MDR-TB treatment. There is no meaningful advantage in using two specimens for the monitoring of MDR-TB patients. A single specimen policy could be safely implemented with negligible clinical effect on MDR-TB patients and favourable resource implications for RNTCP. In India, the estimated incidence of multi-drug resistant (MDR) tuberculosis (TB) cases in 2009 was 99,000. In accordance with international and national guidelines, the progress of each patient during MDR-TB treatment is monitored by sputum culture at a Culture and Drug susceptibility testing (C&DST) laboratory accredited by the national programme. The purpose of follow-up culture monitoring is to assess the response to MDR-TB therapy, in order to guide treatment adjustments. A positive culture indicates poor response to therapy; conversely consistently negative cultures indicate good response. Although collection of two sputum specimens to maximize culture recovery has been recommended for the diagnosis of MDR-TB cases and in international guidelines for the surveillance of drug-resistant tuberculosis, international guidelines are silent as to the number of sputum specimens recommended for follow-up culture examinations in MDR-TB RNTCP in national PMDT guidelines has chosen to collect two specimens for both diagnosis of MDR-TB and for follow-up culture monitoring. The schedule for monitoring involves examining two sputum specimens at months 3, 4, 5, 6, 7, 9, 12, 15, 18, 21, 24 (11 monitoring follow-ups)  [5]. TheA single-specimen policy for follow-up culture examinations would be expected to reduce patient inconvenience, the burden of specimens requiring processing in laboratories and costs. This in turn would mean more available bandwidth for the processing of diagnostic specimens for the detection of MDR-TB. Despite these broad ramifications, there was no information to guide this policy decision. We conducted a retrospective study at three MDR-TB treatment sites in India to determine the incremental yield of a second sputum specimen for culture during follow-up examinations of MDR-TB patients registered during 2008–09 and to determine the impact of changing from two to a single specimen follow-up strategy on case management.This is a cross sectional study involving review of programme records and registers.Under India’s RNTCP, the diagnosis of MDR-TB is made by doing Culture and Drug Susceptibility Testing (C&DST) on patients who fail RNTCP treatment regimens. The diagnosed MDR-TB patients undergo pre-treatment evaluation before starting MDR-TB treatment at MDR-TB treatment sites where the patient is registered, started on treatment and monitored in hospital for a period of at least seven days before referral to domiciliary treatment.th and 5th month culture results are negative. If either or both 4th–5th month culture results remain positive, treatment is extended by one month. Extension of IP beyond 1 month will be decided on the results of sputum culture of 5th or 6th and 6th or 7th months. If the 4th month culture is still awaited after 6 months of treatment, the IP is extended until the result is available, with further treatment being decided according to the culture results. The IP can be extended up to a maximum of 3 months after which the patient will be initiated on CP irrespective of the culture result. The recommended duration of CP is 18 months.Treatment is given in two phases, Intensive (IP) and continuation (CP). The total duration of treatment for regimen of MDR-TB is 24–27 months, depending on the IP duration which should be given for at least six months. Patients will be considered culture converted after having two consecutive negative cultures taken at least one month apart in IP. After six months of treatment, patients are reviewed and treatment changed to CP if both the 4rd to 7th month of treatment  and at 3 monthly intervals from the 9th month onwards till the completion of treatment . All the specimens were subjected to solid culture. During the study period, the C & DST laboratories of these study sites were accredited by RNTCP for solid culture technology. The WHO standardised treatment outcome definitions are used in RNTCP.For follow-up examinations, the specimens are collected and transported under cold chain conditions from the respective District TB Centre and examined by smear and culture at least 30 days apart from the 3All MDR-TB patients registered at three MDR-TB treatment sites during 2008–09, and whose treatment outcome were reported by end of 2011 were included in the study. The three sites were Hyderabad (Andhra Pradesh), Ahmedabad (Gujarat) and Jaipur (Rajasthan).Variables collected for each patient were patient name, DOTS-plus TB number, age, sex, HIV status, date of registration, month of follow-up, date and results of smear examination, date and results of culture examination and treatment outcome. The outcome variables were all culture results. Data were collected into a structured proforma by the trained staff at MDR-TB treatment sites and laboratories. The source of data was from DOTS-plus TB register and C&DST laboratory register.All the data was entered twice by independent data entry operators trained for the purpose into an EpiData database. The two databases were then compared for discrepancies and a final database was created after correcting discrepancies by referring the original records. This final database was then analyzed using EpiData analysis software. Data were cross-tabulated and the following were calculated: the proportion of follow-up examinations which were culture positive, the proportion who were positive on the second specimen only when the first specimen was non positive , and the its inverse (1/X) was determined to calculate the number of second sputum culture examinations, that were done to get one additional culture positive case.The longitudinal culture results of the MDR-TB patients were then examined to estimate hypothetically, the impact of using the results of single (first) specimen only on case management of MDR-TB patients; specifically to find the number of patients who would have missed IP extension, number who would have delayed submission of specimens for second line drug susceptibility testing and number who would have a different outcome declared or led to a delay in declaration of outcome.As this study was a record review of routinely collected RNTCP data, informed consent of individual patients was deemed unnecessary. The same was reviewed and approved by Central TB Division, Ministry of Health and Family Welfare, India andthe Ethics Advisory Group of International Union Against TB and Lung Disease (The Union).A total of 220 MDR-TB patients registered at the three MDR-TB treatment sites were included in the study. During their treatment period 1763 pairs of follow-up culture examinations were done. Data of 19 first culture examination results and 23 second culture examination results were not recorded and hence results of remaining 1721 (98%) pairs were included in the analysis.Of 1721 pairs of follow-up sputum culture examinations done among 220 MDR-TB patients, 451(26%) were positive with either of the two specimens; 29(1.7%) were culture positive only on the second specimen indicating the incremental yield.  To detecAs shown in This is the first study describing the incremental yield of the second sputum specimen during follow-up culture examinations of MDR-TB patients and the impact on case management on changing from two to single specimen follow-up strategy. The strength of the study is that routine programme data was used from the cohort of MDR-TB patients registered under RNTCP, thus reflecting the operational reality. We have adhered to STROBE guidelines on reporting These findings have the following programmatic implications. First, the incremental yield of examining a second specimen was very low at 1.7%, and it is pragmatically redundant to process nearly 60 follow-up specimens to get one additional positive culture. In resource limited settings with limited laboratory capacity, reducing to single specimen follow-up will decrease the laboratory workload considerably and helps in prioritizing the effort towards examining more diagnostic specimens The effect of the single specimen follow-up strategy was limited to a handful of missed positive cultures which in turn would have had very little effect on MDR-TB case management. Effectively, the collection transportation and laboratory processing of 1721 second follow-up cultures led to 3 instances where the intensive phase of treatment is extended. Reassuringly, there were no observed lost opportunities for the detection of treatment failure and referral for second line drug susceptibility testing. Rather, any possible events would have been triggered by an earlier or a later follow-up culture examination. With serial cultures, such events would be likely caught at the next or preceeding follow-up cultures. This is very convincing evidence to move towards single specimen follow-up culture examination and urge RNTCP to consider policy change in this regard.Though, we did not find any instance in this cohort of MDR-TB patients, the other possible impact on case management of a missed positive culture is that we miss the opportunity to identify treatment failure early and initiate steps to examine if the patient has extensively drug resistant tuberculosis. This possibility can be greatly minimized if national programme takes a policy decision to examine all the specimens for second line DST right at the beginning of treatment. However, the current capacity of RNTCP for second line drug susceptibility testing is limited and needs to be strengthened further.This study had some limitations. As in any record–based study, it is limited by the retrospective nature of the data, which may have been inaccurate or incomplete; however, routine supervision and monitoring within the programme is likely to have reduced this. In conclusion, there is no meaningful advantage in using two specimens for the monitoring of MDR-TB patients. A single specimen policy could be safely implemented with negligible clinical effect on MDR-TB patients and favourable resource implications for RNTCP.


Saccharomyces cerevisiae genome, such as the rosette conformation of the genome, the clustering of centromeres, as well as tRNAs, and telomeres. We also demonstrate that simple architectural features of genomic interaction networks, such as cliques, can give meaningful insight into the functional role of the spatial organization of the genome. We show that there is a correlation between inter-chromosomal clique size and replication timing, as well as cohesin enrichment. Together, our network-based approach represents an effective and intuitive framework for interpreting high-throughput genomic interaction data. Importantly, there is a great potential for this strategy, given the rich literature and extensive set of existing tools in the field of network analysis.We propose a network-based approach for surmising the spatial organization of genomes from high-throughput interaction data. Our strategy is based on methods for inferring architectural features of networks. Specifically, we employ a community detection algorithm to partition networks of genomic interactions. These community partitions represent an intuitive interpretation of genomic organization from interaction data. Furthermore, they are able to recapitulate known aspects of the spatial organization of the  The non-random spatial organization of chromosomes in the eukaryotic nucleus is strongly associated with various types of genomic regulation. Spatial compartmentalization has been shown, in many organisms, to correspond to transcriptional regulation, DNA replication, and chromatin states Chromosome conformation capture (3C) technology has enabled the identification of long-range interactions between genomic loci Genomic interaction data is essentially composed of pairwise relationships between genomic regions. Since networks abstractly represent pairwise relationships between objects, this type of data has an inherent network structure. Thus, networks can be used to generate highly intuitive representations of this type of data. Networks are also a convenient and highly flexible framework for storing, analyzing, and integrating interaction data. Furthermore, information of biological interest that is contained in interaction data, such as compartmental characteristics, can be extracted by analyzing the architectural properties of an interaction network. As is necessary to analyze genome-scale datasets, efficient algorithms have been developed to identify some of these network properties in very large networks Here we demonstrate how intuitive, biologically meaningful analyses of large genomic interaction datasets can be achieved purely through network abstractions. Although some groups have begun to employ networks for analyzing gene-gene and other types of interactions from Hi-C data Saccharomyces cerevisiae by Duan et al.In this study, we generate and analyze network models constructed from an unbiased genome-wide chromatin-chromatin interaction dataset generated in et al.The chromosome interaction data was generated by Duan Replication timing data was obtained from McCune et al., Supplemental data 1 et al.ChIP-seq data for cohesin  and cohesin loader  subunits were obtained from Hu et al. Processed gene expression for ORFs were obtained from Nagalakshmi Gene annotations were obtained from the “SGD Genes” track of the UCSC Genome Browser database . Centromere, telomere, and tRNA annotations were obtained from the “SGD Other” track of the UCSC Genome Browser database  Saccharomyces cerevisiae genome were lifted over to sacCer3 using the UCSC Genome Browser liftOver tool All coordinate-based datasets that did not correspond to the sacCer3 assembly of the The networks were built using the NetworkX Python module Clique detection was performed using the find_cliques function in NetworkX. Each node was assigned a maximum clique size, which is the size of the largest clique to which the node belongs. An in-house implementation of the Louvain algorithm was used to perform community detection Enrichments for protein binding sites and genomic features  were calculated using the two-tailed Fisher's exact test. The categories for the contingency table used to calculate the result of the test were, fragments that contain a given feature, and fragments that belong to a given community. Thus, the test calculates the probability that fragment feature assignment and fragment community assignment are independent. In the case of the transcriptional regulators, the FDR was calculated by applying the Benjamini-Hochberg procedure in vivoWe created network models of genomic interactions, where nodes represent genomic loci, and edges represent statistically significant interactions between loci , unless otherwise stated). Several groups have noted the highly stochastic nature of these interactions The known functions of cohesin make it an excellent candidate for a mediator of stable inter-chromosomal interactions. In budding yeast, cohesin has a well established role in mediating inter-chromosomal cohesion between newly replicated sister chromatids In the inter-chromosomal network, we calculated the maximum clique size for each genomic fragment, which is the size of the largest clique of which a given fragment is a member . At eachCohesin has also been shown to be recruited to sites of active replication in budding yeast The clear trends between inter-chromosomal clique size, cohesin enrichment, and replication timing demonstrate that biologically relevant information can be gleaned from the structural properties of genomic interaction networks. Relatively complex information about the interaction behavior of a genomic region can be obtained through simple characterizations of an interaction network. Since clique size is a relatively simple aspect of the inter-chromosomal network architecture, these findings demonstrate the potential for more sophisticated network analyses.Communities are groups of densely connected nodes in a network. In a genomic interaction network, communities represent dense clusters of interacting genomic loci, e.g., chromosome territories. Therefore, the community structure of the genomic interaction network is of great interest, since it reflects how the genome is spatially compartmentalized. The budding yeast genome has been shown to have some degree of compartmentalization, including the clustering of the rDNA locus on chromosome XII ijA is the adjacency matrix of the network , m is the sum of the edge weights in the network, ik is the sum of the edge weights attached to node i, ic is the community to which node i belongs, and δ is the Kronecker delta. Many community detection procedures take the approach of attempting to maximize modularity. Though there is no single rigorous definition of a network community, modularity optimization is widely used to define the community partition of a network. However, optimizing modularity has been shown to be an NP-complete problem, and is thus computationally intractable Detecting communities involves partitioning the network so that nodes within communities are densely connected, and nodes between communities are sparsely connected. A commonly used metric for the quality of a partition is its modularity given by To detect communities in our genomic interaction network, we implemented the so-called Louvain algorithm One concern relevant to community detection in interaction networks is the influence of the significance threshold selected for the inclusion of edges in the network. The selected threshold has a strong influence on the number of edges included in the network, and since the modularity resolution limit is a function of the total number of edges in the network, the inclusion of “noisy” edges in the network increases the minimum detectable community size The inter-chromosomal-only networks had 31,832 and 13,537 edges at the 1% and 0.01% FDR thresholds, respectively. The mean community recapitulation rate for these networks was 84% across the communities in the smaller network. The complete interaction networks had 59,132 and 31,426 edges at the 1% and 0.01% FDR thresholds, respectively. This pair yielded a mean community recapitulation of 93%. These calculations suggest that, at a coarse-grained scale, community detection is highly robust to the selection of significance thresholds for network edges. The subsequent analyses are done on networks with a 1% FDR edge threshold. We selected this less stringent threshold in order to incorporate larger portions of the genome.The inter-chromosomal network contains 2955 nodes and 31,832 edges. The partition solution to this network has three hierarchical levels see . Level 2We looked at enrichment of several chromosomal features and transcriptional regulators in each of the three high-level communities. Community 1 contains all of the centromeres, so not surprisingly it has a highly significant enrichment for centromeres (p = 9.36e-14). It also has a significant enrichment for tRNAs (p = 0.008), which is consistent with the observation of a centromere-proximal spatial cluster of tRNAs The centromere-distal communities had less dramatic enrichments. Community 0 does not contain enrichments for the chromosomal features, or any of the transcriptional regulators we assessed. This is not surprising, considering this community accounts for over half of the genome, and is the most sparsely connected of the three. Although, community 0 tends to be more centromere-distal than community 6, community 6 contains a significant enrichment for telomeres (p = 0.0088). This suggests possible looping associations between telomeres and telomere-distal regions of chromosomes. The size of communities 0 and 6 contribute to their non-specificity; that is, they are low-resolution communities. Therefore, we sought to explore the hierarchical community structure of the genome.Communities at each successive hierarchical level of the detection algorithm represent aggregations of communities in the preceding level. Therefore, the intermediate partitions of the inter-chromosomal network, should give information about the hierarchical structure of the network. However, the partition levels of this network give little indication of hierarchical structure. At level 1, there are three communities nearly identical to the three communities in level 2 . In ordeOne possibility for the lack of evidence for hierarchical structure in the inter-chromosomal network is that the intermediate solutions to the detection algorithm do not have the ability to resolve subcommunities. Moreover, force-directed layouts of the large network may not impose a geometry that allows visual discernment of community structure, especially for subtle communities. To further investigate the possibility of hierarchical structure in the inter-chromosomal network, we performed community detection on the three subnetworks that represent each of the three major communities in the inter-chromosomal network. We treated each of these subnetworks as independent networks. In this section, we will only refer to the communities generated by the final partition of these subnetworks. Also, these communities-within-communities will henceforth be referred to as “subcommunities”. To assess the presence of hierarchical community structure in this data, we looked for evidence of modular structure in the subnetworks, and biological meaning in the subcommunities.Since partitions of random networks can have highly variable modularity, a modularity value on its own does not have a meaning The subnetwork induced by community 1 (the centromeric community) of the inter-chromosomal network had the greatest degree of modularity over random . This isCommunity 6 of the inter-chromosomal network showed the second largest modularity over random (Z = 7.63). Like the partition of community 1, we see large contiguous chromosomal segments assigned to a single subcommunity . HoweverInter-chromosomal community 0 shows the weakest modularity over random (Z = 5.9). Accordingly, it has very few large contiguous subcommunities . The onlNext, we partitioned the network containing both intra- and inter-chromosomal edges (which will be referred to as the “complete network”) into communities. The interpretation of this network has a major caveat associated with it. The FDRs of intra-chromosomal and inter-chromosomal links were calculated using different assumptions and probability models  Scc2 and (B) Scc4 with respect to maximal fragment clique size. Like cohesin itself, cohesin loader enrichment increases with number of interacting chromosomes.(TIF)Click here for additional data file.Figure S2Cohesin enrichment vs. intra-chromosomal maximal clique size. Enrichment of cohesin subunits (A) Smc3 and (B) Scc1 with respect to maximal fragment clique size in the intra-chromosomal network. This plot includes intra-chromosomal cliques across all chromosomes. Unlike the inter-chromosomal cliques, cohesin enrichment and intra-chromosomal clique size are independent.(TIF)Click here for additional data file.Figure S3Cohesin loader enrichment vs. intra-chromosomal maximal clique size. Enrichment of cohesin loader subunits (A) Scc2 and (B) Scc4 with respect to maximal fragment clique size in the intra-chromosomal network. Like cohesin itself, cohesin loader enrichment and intra-chromosomal clique size are independent.(TIF)Click here for additional data file.Figure S4Replication timing vs. intra-chromosomal maximal clique size. Unlike inter-chromosomal cliques, intra-chromosomal clique size and replication timing are independent.(TIF)Click here for additional data file.Figure S5Expression vs. inter- and intra-chromosomal maximal clique size. Gene expression level is independent of the (A) inter-chromosomal and (B) intra-chromosomal clique size of its genomic locus.(TIF)Click here for additional data file.Figure S6Intermediate solutions to community detection in the inter-chromosomal network. Scaled chromosomes, centered on centromeres. (A) The level 1 partition of the inter-chromosomal network is similar to the level 2 partition Click here for additional data file.Figure S7Partitions of inter-chromosomal community 6 and 0. (A) The partition of the subnetwork induced by community 6 shows several large continuous community assignments, indicating some modular community structure. (B) The partition of the community 0 subnetwork is highly fragmented, indicating very little modular community structure.(TIF)Click here for additional data file.Figure S8Level 0 partition of the complete network. The level 0 partition of the network containing both inter- and intra-chromosomal interactions shows very similar community structure to the level 1  partition. This indicates that there is little hierarchical community structure information in the intermediate solution to the final partition.(TIF)Click here for additional data file.Table S1Inter-chromosomal network maximum clique size. Table of genomic fragments and the size of the largest inter-chromosomal clique to which each fragment belongs.(XLSX)Click here for additional data file.Table S2Inter-chromosomal network community assignments. Table of genomic fragments and the community numbers to which they belong. Community numbers are given for each level of the partition.(XLSX)Click here for additional data file.Table S3Inter-chromosomal community 1, subcommunity assignments. Table of genomic fragments in inter-chromosomal community 1, and the subcommunity numbers to which they belong. Subcommunity numbers are given for each level of the partition.(XLSX)Click here for additional data file.Table S4Inter-chromosomal community 6, subcommunity assignments. Table of genomic fragments in inter-chromosomal community 6, and the subcommunity numbers to which they belong. Subcommunity numbers are given for each level of the partition.(XLSX)Click here for additional data file.Table S5Inter-chromosomal community 0, subcommunity assignments. Table of genomic fragments in inter-chromosomal community 0, and the subcommunity number to which they belong. Subcommunity numbers are given for each level of the partition.(XLSX)Click here for additional data file.Table S6Inter-chromosomal community 0, subcommunity 25 TF enrichments. Transcription factors enriched in inter-chromosomal community, subcommunity 25 (FDR <1%).(XLSX)Click here for additional data file.Table S7Complete network community assignments. Table of genomic fragments and the community numbers to which they belong. Community numbers are given for each level of the partition.(XLSX)Click here for additional data file.


Patients with ST-elevation myocardial infarction (STEMI) not treated with primary or rescue percutaneous coronary intervention (PCI) are at risk for recurrent ischemia, especially when viability in the infarct-area is present. Therefore, an invasive strategy with PCI of the infarct-related coronary artery in patients with viability would reduce the occurrence of a composite end point of death, reinfarction, or unstable angina (UA).Patients admitted with an (sub)acute myocardial infarction, who were not treated by primary or rescue PCI, and who were stable during the first 48 hours after the acute event, were screened for the study. Eventually, we randomly assigned 216 patients with viability (demonstrated with low-dose dobutamine echocardiography) to an invasive or a conservative strategy. In the invasive strategy stenting of the infarct-related coronary artery was intended with abciximab as adjunct treatment. Seventy-five (75) patients without viability served as registry group. The primary endpoint was the composite of death from any cause, recurrent myocardial infarction (MI) and unstable angina at one year. As secondary endpoint the need for (repeat) revascularization procedures and anginal status were recorded.The primary combined endpoint of death, recurrent MI and unstable angina was 7.5% (8/106) in the invasive group and 17.3% (19/110) in the conservative group . During follow up revascularization-procedures were performed in 6.6% (7/106) in the invasive group and 31.8% (35/110) in the conservative group . A low rate of recurrent ischemia was found in the non-viable group (5.4%) in comparison to the viable-conservative group (14.5%). .We demonstrated that after acute MI (treated with thrombolysis or without reperfusion therapy) patients with viability in the infarct-area benefit from a strategy of early in-hospital stenting of the infarct-related coronary artery. This treatment results in a long-term uneventful clinical course. The study confirmed the low risk of recurrent ischemia in patients without viability.NCT00149591.ClinicalTrials.gov:   Management of acute myocardial infarction (AMI) has changed considerably over the last two decades -3. OptimAfter successful thrombolysis, more than 50% of patients have a significant residual stenosis and about 20-30% suffer from recurrent ischemic events because of plaque-instability in the infarct-related coronary artery -13. ViabIn contrast, the cardiac event-rate in patients without viability was low and did not change by an invasive strategy. In current clinical practice, viability-testing is not used as a tool for post-myocardial infarction risk-assessment and patient management. We report the results of the Viability-guided Angioplasty after acute Myocardial Infarction (VIAMI) trial, which tested the hypothesis that a strategy of viability guided angioplasty with stenting after AMI in patients treated with thrombolysis or who were too late for reperfusion therapy and remained stable for 48 hours, would reduce the occurrence of a composite end point of death, reinfarction, or unstable angina.The methods used in the trial have been described previously . In brieStable patients revealed no signs of ongoing ischemia based on electrocardiographic characteristics or persistent chest discomfort. Patients who were admitted within 6 hours after symptom onset, received thrombolysis combined with heparin. Patients admitted more than 6 hours after symptom onset, received only heparin or low weight molecular heparin (LWMH).Patients < 80 years of age were considered suitable for the study when they met the criteria for definite myocardial infarction, i.e. an significant rise in creatine kinase-MB levels , 1 mm ST segment elevation in two or more standard leads or 2 mm ST segment elevation in two contiguous chest leads, and/or the development of Q waves.Patients underwent low dose dobutamine echocardiography (LDDE) for the detection of viability within 72 hours after AMI. It is a safe and well-validated bedside test with a diagnostic accuracy of about 80%, which is comparable to scintigraphical techniques (SPECT/PET) . Before Viability was defined as the improvement of wall motion abnormalities (WMA's) in two or more segments of the infarct zone. Patients without WMA's were not included in this trial. In case of poor acoustic window ultrasound contrast agents were used to improve image quality and diagnostic yield.All images were sent to the core-lab  and were analyzed by 2 experienced observers. A third observer was used in case of disagreement to reach consensus. All eligible patients provided written informed consent. The study complied with the Declaration of Helsinki and all ethics committees of the participating centers approved the protocol.Patients with viability in the infarct-area were randomized to an invasive or a conservative treatment strategy. Permuted block randomization was performed with a block size of ten. All patients were treated with aspirin, beta blockers, angiotensin-converting-enzyme inhibitors, statins as accepted by international guidelines ,2.The invasive strategy patients underwent in-hospital coronary angiography with the intention to perform PCI with stenting of the infarct-related coronary artery (IRA). In the conservative group an ischemia-guided approach was adopted with stress testing before hospital discharge. After a positive test for ischemia coronary angiography was strongly recommended. If an intervention was performed, it was considered planned and not interpreted as an event. Patients without viability served as a registry group also with a long-term follow-up.Angiography and PCI was performed as soon as possible after randomization. When a significant (≥ 50%) stenosis or occlusion of the infarct-related coronary artery was found, PCI with stenting was performed when feasible. In all cases where PCI was performed, abciximab was used according to the EPILOG protocol . A bolusThe primary endpoint was the composite of death from any cause, recurrent infarction and unstable angina at 1-year follow-up. Secondary endpoints were the need for revascularization and the occurrence of angina pectoris (Canadian Cardiovascular Society classification (CCS)).A recurrent myocardial infarction was diagnosed if there was an increase in the total creatine kinase and MB isoenzyme activity (2 times ULN) and either a history of ischemic chest discomfort or electrocardiographic changes indicative for trans mural ischemia or necrosis. Reinfarction during hospitalization required a decrease of cardiac enzymes, followed by a subsequent rise to a level of 2 times ULN and 50% above a previous measured value.For the diagnosis of unstable angina, patients had to be rehospitalized with ischemic chest pain or discomfort occurring at rest or with minimal exertion. In addition, the need for intravenous medical intervention and/or objective evidence of myocardial ischemia was required.Members of an independent clinical event and end point committee, who were unaware of the treatment assignments of the patients, adjudicated all end points.The VIAMI-trial was conducted to investigate the differences in clinical outcome between an invasive and a conservative strategy in patients with demonstrated viability in the infarct-area. The expected event rate in the viability positive group was estimated to be 35 percent. To demonstrate with a power of 80%  that PCI leads to a 50% reduction in event rate in the invasive group compared to the conservative group, 200 patients would be needed in each group.As a formal stopping rule for the study the following was used: if one of the treatment strategies appeared significantly superior at interim analysis (p ≤ 0.01), the study would be stopped. Interim analysis was performed each time another 100 patients were included.Baseline descriptive data are presented as a mean ± standard deviations (SD). Differences in clinical and echocardiographic variables are assessed by unpaired Student's t-test. Differences between proportions are assessed by chi-square analysis; a Fisher's exact test is used when appropriate. Event-free survival curves are computed with the Kaplan-Meier method, and the differences between these curves are tested with a log-rank test. The Cox proportional hazards regression analysis was used to estimate the treatment effect as hazard ratio (HR) with 95% confidence intervals. Besides the "crude" effects, adjustments were made for DM, hypertension, hypercholesterolemia, current smoking, family history of CAD (model a), clinical history  and medication use at baseline  (model b) and for all covariates (model c).All analyses were performed on an intention-to-treat basis. Outcome per-protocol was also evaluated, since this would reflect the true influence of PCI on clinical outcome. Because after randomization there was a median waiting-time of two days before a revascularization procedure was performed inevitably some events occurred. In the per-protocol analysis these events are excluded from analysis, because they occurred before the by protocol demanded intervention. To make a fair comparison between the two groups in the per-protocol analysis we also excluded the events in the conservative group occurring during the first two days after randomization. All analyses were performed with the use of SPSS software, version 16.0 .Between April 2001 and January 2006, 216 patients were enrolled in the trial. Of these, 106 patients were randomly assigned to the invasive strategy and 110 patients to the conservative strategy. The pre-specified number of patients was not achieved. This was mainly caused by a slowing down of inclusion-rate in the second half of the study period due to the fast introduction of widely available primary PCI in the Netherlands (> 90%). At the start of this study, 50% of all AMI were still being treated with thrombolysis in the Netherlands.The baseline characteristics of the patients in the two randomized groups were similar except for a lower prevalence of statin use in the group assigned to the conservative strategy (p = 0.02)Table . The meaThe primary endpoint  occurred in 27 patients (8 in the invasive group and 19 in the conservative group). By intention-to-treat analysis the one-year event-free survival was 92.5% percent in the invasive strategy group and 82.7% in the conservative strategy group Figure .The Kaplan-Meier curves of the adjusted per-protocol analysis revealed an estimated one-year event free survival of 95.3 percent in the invasive group and 85.5 percent in the conservative group Figure .There was no difference in event-rate between patients who were treated with thrombolysis and patients who did not receive reperfusion therapy. Primary endpoint events were similar in these two groups  Figure .Patients in the invasive group had a significantly lower anginal class (CCS) (p = 0.021)Table . After dThere was no difference in revascularization procedures performed in patients treated with thrombolysis and in those who received no reperfusion therapy Figure .No significant difference in the primary composite endpoint was found in the comparison between the non-viable and the viable-conservative group Figure . In the The VIAMI-trial is the first RCT investigating a viability-guided invasive approach in patients who were at least 48 hours stable after acute MI (not treated with primary PCI), demonstrating that only patients with viable tissue in the infarct-area showed benefit from an early in-hospital culprit vessel revascularization.Viability was used as a sensitive marker of risk of recurrent ischemia and recurrent infarction, with the notion that recurrent ischemic events in the post-MI period in most instances is related to re-thrombosis in the infarct-related coronary artery, combined with residual viable tissue in the infarct-area.Several studies evaluated early invasive intervention after thrombolysis -25. The With regard to the recommended treatment post-thrombolysis, the AHA/ACC and ESC guidelines differ diminutively. The AHA/ACC guidelines recommend an early pharmacoinvasive strategy after thrombolysis in high risk patients . In non-high risk patients such a strategy could be considered, especially if ischemic symptoms persist and failure to reperfuse is suspected . The ESC guidelines and ESC guidelines for PCI recommend a routine invasive strategy within 24 hours after successful thrombolysis if available . In stable patients who did not receive reperfusion therapy, angiography could be considered before discharge  ,2,27. HoAs an unexpected finding, patients without viability underwent as many revascularization procedures as patients with viability who were randomized to the conservative strategy. Although patients without viability had a low recurrence-rate of acute coronary syndrome, they had similar reported CCS-class of angina compared to the viable-conservative patients. Despite a lower rate of spontaneous ischemic events and less proven ischemia during stress-testing, treating physicians decided to refer patients for an invasive procedure quite as often in the non-viable patient group. The explanation for this remains speculative.The results of the VIAMI trial suggest that routine angioplasty in the early post-MI period in stable patients (whether or not treated with thrombolysis) is not mandatory. Especially in patients were the transport time for primary PCI is largely exceeding 90 minutes as recommended by the AHA/ACC guidelines, thrombolysis could be an interesting alternative with the VIAMI-trial approach. Also in patients presenting too late for reperfusion therapy (> 12 hours). Early viability-testing with low-dose dobutamine echocardiography provides us with a tool to identify high-risk patients who will have clinical benefit from a revascularization procedure later on.The applicability of LDDE in daily practice is somewhat limited because about 10-15% of patients have poor acoustic windows. In our study this problem could partially be overcome with selective use of ultrasound contrast agents.The pre-specified number of patients was not achieved. This was mainly caused by a slowing down of inclusion-rate in the second half of the study period due to the fast introduction of widely available primary PCI in the Netherlands. Although the primary event-rate in the viable-conservative group was lower than expected (17.3%) the relative risk-reduction was higher than expected (56.6%). This indicates an improvement over time in the pharmacological post-MI treatment . Also, it implicates further improvement in the efficacy and safety-profile of revascularization procedures. During the inclusion-period of the VIAMI-trial treatment with clopidogrel was not standard care in patients without stents. Standard treatment with clopidogrel according to current guidelines [to GV and RvL]. GV received research grants from Eli Lilly, Boehringer Ingelheim Pharmaceuticals, and Bristol-Myers Squibb.RvL contributed to acquisition of data, analysis and interpretation of data, drafting the manuscript, and statistical analysis. GV contributed to this work in conception and design, interpretation of data, drafting of the manuscript, and supervision.LB contributed to critical revision, and technical support. OK, JB, and AvR contributed to conception, design, and supervision. JT contributed to conception and design, statistical analysis, and supervision. All authors read and approved the final manuscript.


Biomarkers of peripheral muscle fatigue (BPMFs) are used to offer insights into mechanisms of exhaustion during exercise in order to detect abnormal fatigue or to detect defective metabolic pathways. This review aims at describing recent advances and future perspectives concerning the most important biomarkers of muscle fatigue during exercise.BPMFs are classified according to the mechanism of fatigue related to adenosine-triphosphate-metabolism, acidosis, or oxidative-metabolism. Muscle fatigue is also related to an immunological response. impaired calcium handling, disturbances in bioenergetic pathways, and genetic responses. The immunological and genetic response may make the muscle susceptible to fatigue but may not directly cause muscle fatigue. Production of BPMFs is predominantly dependent on the type of exercise. BPMFs need to change as a function of the process being monitored, be stable without appreciable diurnal variations, correlate well with exercise intensity, and be present in detectable amounts in easily accessible biological fluids. The most well-known BPMFs are serum lactate and interleukin-6. The most widely applied clinical application is screening for defective oxidative metabolism in mitochondrial disorders by means of the lactate stress test. The clinical relevance of most other BPMFs, however, is under debate, since they often depend on age, gender, physical fitness, the energy supply during exercise, the type of exercise needed to produce the BPMF, and whether healthy or diseased subjects are investigated.Though the role of BPMFs during fatigue is poorly understood, measuring BPMFs under specific, standardised conditions appears to be helpful for assessing biological states or processes during exercise and fatigue. Fatigue in response to exercise (exercise (-induced) fatigue) can be caused by mental disorders, organic central nervous system (CNS) abnormalities , or by peripheral nervous system (PNS) dysfunction or skeletal muscle disease ,2. FactoNeurological causes include:Central fatigueMental disordersOrganic CNS abnormalitiesPeripheral fatiguePNS Skeletal muscle diseases Non-neurological causes include:Cardiac disordersPulmonary disorders (e.g. chronic obstructive pulmonary disease (COPD)Hematologic disordersMetabolic diseasesRenal problemsMalignanciesOvertraining syndromeChronic fatigue syndrome (CFS)Exercise-induced muscle fatigue has to be clearly delineated from fatigability in the resting stage (fatigue prior to exercise), which may be caused by similar or other factors than exercise-induced muscle fatigue. Exercise-induced muscle fatigue is defined as a reversible loss of muscle force (muscle contractility) during work over time ,5. CliniVoluntary muscle exercise can be carried out in an incremental or constant manner (endurance), or in a dynamic  or static mode. Exercise can also be exhaustive or non-exhaustive. Exhaustive exercise includes acute, very prolonged exercise, a period of intensified training, non-functional overreaching and the overtraining syndrome. Exercise can be carried out under aerobic or anaerobic conditions. Examples of partially anaerobic exercise include sprinting, where maximal efforts last <1 minute, and exercise under high-force isometric conditions. Only a very few types of exercise are entirely anaerobic, and these necessarily last for only a few seconds. Contractions during exercise may be isometric or non-isometric, isokinetic (constant velocity throughout movement) or non-isokinetic (changing velocity during contraction), or concentric (muscle shortening) or eccentric (muscle lengthening). All of these different conditions are not exclusive, i.e. a cycling exercise may be constant, dynamic, non-isometric and concentric.Muscle fatigue is defined as decreased ability to generate appropriate amounts of muscle force or power during on-going contractile activity. Muscle fatigue may vary according to the cause or underlying mechanisms, with variable rates of recovery. For example, fatigue brought on by a marathon is different from fatigue that prevents a 10th bench press repetition at a given resistance. Exercise-induced muscle fatigue can occur shortly after onset of exercise (acute muscle fatigue) or after a constant, high-intensity exercise had been carried out for a prolonged period of time (delayed exercise-induced fatigue), which is characterized by tiredness only after a longer duration of constant exercise. Acute muscle fatigue is usually maximal immediately post-exercise. Exercise-induced muscle fatigue can develop under a sub-maximal as well as maximal intensity exercise. Exercise-induced fatigue can occur in healthy or diseased subjects, and depends on age, gender, physical fitness, mode and duration of exercise, and the underlying disease.Contrary to fatigue, the term fatigability is defined as rate of loss of muscle force over time. Fatigability is an interactive determinant of the state of fatigue versus rested. Only few studies have addressed the relation between fatigability and biomarkers of muscle fatigue. Muscle fatigue has to be delineated from the sensation of tiredness. Muscle fatigue that develops during submaximal aerobic exercise is not necessarily reflected by a sensation of tiredness, but a reduced ability to develop force and power. Recovery from fatigue is defined as the rate at which muscle function returns to baseline after becoming fatigued. Recovery represents an important marker of fatigue severity and biomarker responsiveness. Like fatigability, recovery represents an interactive determinant of the state of fatigue versus rested. Both, fatigue severity and recovery are influenced by fatigability and the degree to which muscle dysfunction is provoked.Decrease in force production during exercise can be regarded as a safety mechanism. If fatigue would not occur or was delayed, structural damage to muscle cells and supportive tissues would occur during the workout. Skeletal muscle is generally composed of slow-twitch and fast-twitch muscle fibres. Slow-twA biomarker  is a measurable product or substance of an organism that is used as an indicator of a biological state to objectively measure physiological or pathogenic processes in the body that occur during health, disease or in response to pharmacological treatment. In mediBiomarkers can be categorized according to the type of exercise or workload carried out ) or accorThe concentration of a biomarker produced depends on the amount of fatigue that is induced and on the type and duration of the exercise. A further classification relies on the biochemical pathway from which the biomarker originates Table.However, the most plausible classification of biomarkers of muscle fatigue follows the mechanisms of fatigue and the metabolic changes during fatigue. Since there is no single cause of muscle fatigue, there is also no single biomarker for assessing muscle fatigue. Studies of several biomarkers during muscle strength decline showed that multiple elevated catabolic biomarkers are a better predictor of muscle fatigue than a single biomarker.Biomarkers of muscle fatigue are different for exercise that lasts 20s in duration, which have an anaerobic energy demand of up to 90%, compared to exercise that lasts 20s to 1 minute, which is maintained by both an anaerobic and aerobic energy supply, and compared to exercise that lasts > 1 minute, for which the energy supply is > 50% aerobic. Biomarkers of muscle fatigue from low- and high-intensity exercise are thus different and their return to normal values depends on the rate of recovery of muscle performance. If fatigue is still present but the biomarker is back to pre-exercise values, then the value of such a biomarker is questionable. A simple way to assess fatigue resistance is to compare the maximal force a muscle (group) can generate after contractile activity with the force/power the muscle could generate before onset of the contractile activity.2+ release and handling[Initially, it was believed that acidosis is the major cause of muscle fatigue. MeanwhiTo which degree shortage in ATP contributes to muscle fatigue is under debate. Though decrease of muscle ATP can be observed during repetitive or prolonged muscle contraction, there are indications that decrease in high energy compounds ) are not enough to result in even a small level of muscle dysfunction. There aAt the beginning of exercise the total adenine nucleotide pool (ATP + ADP + adenosine-monophosphate (AMP)) remains constant, since ADP is reconverted to ATP in order to maintain the delivery of energy for exercise. With inA key role for muscle fatigue plays intracellular acidosis. Even minimal decrease in muscle pH interferes with cross-bridge binding and ATPase activity due to competitive binding and reduced enzyme function. Cross bridge detachment is unaffected by fatigue whereas cross bridge attachment is reduced during fatigue. DecreasIn cases, in which the oxidative phosphorylation of ADP to generate ATP fails to meet the energy requirement of the myocyte, which is the case under acidic conditions, the production of ATP shifts from aerobic processes  to anaerobic glycolysis or glycogenolysis (the processing of glucose/glycogen). WhetherReference limits for serum lactate are reported as > 2 mmol/l. Serum lIf the consumption of ATP exceeds the ATP supply, the ATP:ADP ratio decreases and impairs the functioning of the major ATPases in myocytes, including myosin ATPase (70%), endoplasmatic reticulum Ca-ATPase (25%), and sarcolemmal Na/K-ATPase. ImpairmReference limits of ammonia have been repeatedly reported but depend on a number of variables. Serum ammonia does not seem to be age-dependent. DependeHypoxanthine is a naturally occurring purine derivative, which occasionally is a constituent of nucleic acids, where it is present in the anticodon of tRNAs in the form of inosine. Hypoxanthine and xanthine are derived from the degradation of purinic nucleotides . HypoxanReference limits for hypoxanthine were reported to range between 0 and 8 μmol/l. InfluenOne of the most promising groups of biomarkers of muscle fatigue is that related to oxidative stress, which is characterized by the enhanced production of ROS . ROS havThiobarbituric acid-reactive substances (TBARS) are low-molecular weight end products formed during the decomposition of lipid peroxidation products that react with thiobarbituric acid to form a fluorescent red adduct. TBARS occur in the serum most likely due to peroxidation of low-density lipoproteins and oxygen-mediated injury of myocyte membranes. TBARS aReference limits for serum TBARS at baseline were reported as 6.8-8.0 μM but are in vivo, derive from peroxidation of essential fatty acids (mainly arachnoidic acid) catalysed by ROS and without the action of cyclo-oxygenase. Isoprostanes are esterified to phospholipids in cell membranes and are released in the free form to circulate in body fluids by the action of phospholipases[Isoprostanes are prostaglandin-like compounds that, olipases. Isoprosolipases. IsoprosReference limits of isoprostanes in the serum were reported as 1.5-1.8 ng/ml but depe2max for 30 min and intermittent dumbbell squatting, protein carbonyls increased 1.6-2.4-fold 24 h post-exercise[Protein carbonyls are mainly derived from the oxidation of albumin or other serum proteins. Proteinexercise. After uexercise. The posexercise. In overGlutathione (GSH) is a pseudo-tripeptide that is present in nearly all cells at high concentrations. Glutathione is one of the most important antioxidants and an important reserve of cysteine. GSH is also important for the phase-II biotransformation of toxic substances. For this purpose, halogen-, sulphate-, sulphonate-, phosphate- or nitrate groups are substituted by GSH. In addition to ROS scavenging, GSH is required for the regeneration of ascorbic acid and α-tocopherol. GSH is 2max for 30 min and intermittent dumbbell squatting, GSH decreased by 21% whereas GSSG increased by 25%[Reference limits of GSH in the serum were reported as 0.36-0.41 mM but depend on a number of conditions. GSH levd by 25%. In heald by 25%.2O2) at low levels of exercise, while at higher training volumes the production of H2O2 exceeds the capabilities of GPX. To compensate for the insufficient clearance of H2O2 by GPX, the production of catalase is increased at higher levels of exercise[2O2 is formed by dismutation of superoxide (O2-), which is formed by xanthine-oxidase during the degradation of IMP to uric acid[2O2 causes loss of muscle contractility and thus augments muscle fatigue[Glutathione peroxidase (GPX) is an enzyme that scavenges hydrogen peroxide , metal chelators, and nonspecific antioxidants ). The non-enzymatic antioxidant properties of the TAC are usually measured by means of the ferric reducing antioxidant power (FRAP) reagens. TAC is Among 13 healthy subjects, aged 18-22y, TAC ranged between 1.26 and 1.89 mmol/l. TAC seeIn addition to ATP-depletion and ROS production, exercise and fatigue trigger an inflammatory reaction. After exercise, T-lymphocytes are mobilized into the blood. Available data indicate that contracting skeletal muscle releases myokines (cytokines produced from muscle) to create a systemic anti-inflammatory environment against this inflammatory reaction and to exert specific endocrine effects on visceral fat. This anAerobic physical activity elicits the mobilization of T-lymphocytes into the blood from peripheral lymphoid compartments immediately after exercise, which is quickly followed by lymphocytopenia during the recovery phase. This efReference limits of T-lymphocytes depend on age, sex, and the method applied. T-cellsInterleukin-6 (IL-6) belongs to a group of cytokines that regulate the body’s inflammatory reaction. IL-6 acts as both a pro-inflammatory  and anti-inflammatory (myocytes) cytokine,82. IL-6Among 54 healthy controls, reference limits of IL-6 ranged between 1.0 and 4.8 pg/ml but were sex-related. IL-6 seTNF-α is a pro-inflammatory cytokine that is predominantly produced by macrophages and able to induce apoptosis, inflammation, cell proliferation and cell differentiation, and to inhibit tumourgenesis and viral replication. TNF-α is part of the cytokine system, which modulates the organo-neogenesis. TNF-α indirectly causes insulin resistance by increasing the release of free fatty acids from fat tissue and increasing lipolysis in adipocytes. DysreguReference limits of TNF-α under resting conditions ranged between 0.78 and 3.12 pg/ml and seemExercise results in the upregulation of several genes, as can be demonstrated by increases in mRNA for these genes. For example, the transcriptional rate of the IL-6 gene is markedly enhanced and IL-6 mRNA upregulated during muscle contraction. In rest2O2, vitamin E, and the nonspecific antioxidants albumin and ascorbic acid. In patients with CFS, vitamin E levels in the serum significantly decreased with increasing fatigue[Potential BPMFs due to oxidative stress include H fatigue. Five mi fatigue. Further fatigue. AOPP ha fatigue,18,95.A number of biomarkers turned out unsuitable as a biomarker for muscle fatigue during exercise.Parameters that failed to serve as biomarkers of muscle fatigue so far include elastase, IL-1beta and complement C4a,95. The Although biomarkers appear to be a valuable tool for measuring and monitoring muscle fatigue, it is still under debate, which ones are reliable and the most relevant for clinical use. Among the many biomarkers reported here, the most accurate and valid biomarkers of muscle fatigue are serum lactate and IL-6, but ammonia, oxipurines, the lymphocyte counts or oxidative stress parameters were also investigated for their impact on muscle fatigue. The best biomarkers for monitoring high-intensity muscle fatigue include lactate, IL-6, TBARS, oxipurines and inorganic phosphate. BiomarkBiomarkers are reliable tools for measuring and monitoring muscle fatigue, but the parameters that are the most applicable for this purpose are still being debated. Reasons for the unsatisfactory situation are that validity and reliability, but also proven effectiveness in assessing muscle fatigue, are strongly dependent on the wide range of settings  and on the range of population groups . Other reasons are the still poorly understood physiological changes that skeletal muscle undergoes during fatigue and the huge number of candidates for biomarkers of muscle fatigue, which have either been barely investigated or do not fulfill the criteria of a biomarker. Furthermore, the causal relationship between the biomarkers described and fatigue still needs to be proven, although some correlate substantially with the loss of muscle function. Different assay methods for the discussed biomarkers may also have an impact on their reliability. So far, the most frequently investigated and most widely applied biomarkers of muscle fatigue are serum lactate and IL-6. Additionally, ammonia, leukocytes and oxidative stress parameters are gaining increasing attention. Biomarkers of muscle fatigue could be a prognostic tool for identifying subjects at increased risk of strength decline, but it is unknown whether the interventional lowering of biomarkers can prevent or alleviate muscle strength loss during exercise.There are neither financial nor non-financial competing interests.JF was responsible for the design, literature search, discussion and writing of the text.The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1471-2474/13/218/prepub