Origin of facilitation of motor-evoked potentials after paired magnetic stimulation: direct recording of epidural activity in conscious humans

A magnetic transcranial conditioning stimulus given over the motor cortex at intensities below active threshold for obtaining motor-evoked potentials (MEPs) facilitates EMG responses evoked at rest in hand muscles by a suprathreshold magnetic stimulus given 10-25 ms later. This is known as intracortical facilitation (ICF). We recorded descending volleys produced by single and paired magnetic motor cortex stimulation through high cervical epidural electrodes implanted for pain relief in six conscious patients. At interstimulus intervals (ISIs) of 10 and 15 ms, although MEP was facilitated, there was no change in the amplitude or number of descending volleys. An additional I wave sometimes was observed at 25 ms ISI. In one subject, we also evaluated the effects of reversing the direction of the induced current in the brain. At 10 ms ISI, the facilitation of the MEPs disappeared and was replaced by slight suppression; at 2 ms ISI, there was a pronounced facilitation of epidural volleys. Subsequent experiments on healthy subjects showed that a conditioning stimulus capable of producing ICF of MEPs had no effect on the EMG response evoked by transmastoidal electrical stimulation of corticospinal tract. We conclude that ICF occurs because either 1) the conditioning stimulus has a (thus far undetected) effect on spinal cord excitability that increases its response to the same amplitude test volley or 2) that it can alter the composition (but not the amplitude) of the descending volleys set up by the test stimulus such that a larger proportion of the activity is destined for the target muscle.     

Transcranial magnetic stimulation: new insights into representational cortical plasticity

In the last decade, transcranial magnetic stimulation (TMS) has been used increasingly as a tool to explore the mechanisms and consequences of cortical plasticity in the intact human cortex. Because the spatial accuracy of the technique is limited, we refer to this as plasticity at a regional level. Currently, TMS is used to explore regional reorganization in three different ways. First, it can map changes in the pattern of connectivity within and between different cortical areas or their spinal projections. Important examples of this approach can be found in the work on motor cortex representations following a variety of interventions such as immobilization, skill acquisition, or stroke. Second, TMS can be used to investigate the behavioural relevance of these changes. By applying TMS in its "virtual lesion" mode, it is possible to interfere with cortical function and ask whether plastic reorganization within a distinct cortical area improves function. Third, TMS can be used to promote changes in cortical function. This is achieved by using repetitive TMS (rTMS) to induce short-term functional reorganization in the human cortex. The magnitude and the direction of rTMS-induced plasticity depend on extrinsic factors (i.e. the variables of stimulation such as intensity, frequency, and total number of stimuli) and intrinsic factors (i.e. the functional state of the cortex targeted by rTMS). Since conditioning effects of rTMS are not limited to the stimulated cortex but give rise to functional changes in interconnected cortical areas, rTMS is a suitable tool to investigate plasticity within a distributed functional network. Indeed, the lasting effects of rTMS offer new possibilities to study dynamic aspects of the pathophysiology of a variety of diseases and may have therapeutic potential in some neuropsychiatric disorders. Electronic Publication     

Transmission of human mtDNA heteroplasmy in the Genome of the Netherlands families: support for a variable-size bottleneck

Although previous studies have documented a bottleneck in the transmission of mtDNA genomes from mothers to offspring, several aspects remain unclear, including the size and nature of the bottleneck. Here, we analyze the dynamics of mtDNA heteroplasmy transmission in the Genomes of the Netherlands (GoNL) data, which consists of complete mtDNA genome sequences from 228 trios, eight dizygotic (DZ) twin quartets, and 10 monozygotic (MZ) twin quartets. Using a minor allele frequency (MAF) threshold of 2%, we identified 189 heteroplasmies in the trio mothers, of which 59% were transmitted to offspring, and 159 heteroplasmies in the trio offspring, of which 70% were inherited from the mothers. MZ twin pairs exhibited greater similarity in MAF at heteroplasmic sites than DZ twin pairs, suggesting that the heteroplasmy MAF in the oocyte is the major determinant of the heteroplasmy MAF in the offspring. We used a likelihood method to estimate the effective number of mtDNA genomes transmitted to offspring under different bottleneck models; a variable bottleneck size model provided the best fit to the data, with an estimated mean of nine individual mtDNA genomes transmitted. We also found evidence for negative selection during transmission against novel heteroplasmies (in which the minor allele has never been observed in polymorphism data). These novel heteroplasmies are enhanced for tRNA and rRNA genes, and mutations associated with mtDNA diseases frequently occur in these genes. Our results thus suggest that the female germ line is able to recognize and select against deleterious heteroplasmies.Heteroplasmy (intra-individual variation) in mitochondrial DNA (mtDNA) plays an important role in mtDNA-related diseases and has also been implicated in aging and cancer (Greaves et al. 2012; Wallace 2012; Chinnery and Hudson 2013; Lombès et al. 2014). Most mtDNA mutations that cause diseases due to defects in mitochondrial function exist as heteroplasmies and only cause disease symptoms when the frequency of the mutant allele exceeds a particular threshold (Wallace and Chalkia 2013). Below this threshold, individuals are asymptomatic, presumably because there are sufficient functional mitochondria for normal metabolism. Changes in the frequency of pathogenic mutations during the transmission of heteroplasmies from mothers to offspring can thus play an important role in the disease risk of the offspring. However, most of our knowledge concerning the dynamics of heteroplasmy transmission comes from studies of pathogenic mutations (Monnot et al. 2011; Shen et al. 2012; de Laat et al. 2013; Wallace and Chalkia 2013), which in blood have been shown to decrease over time and hence may not accurately reflect the overall level of such pathogenic mutations within an individual (Poulton and Morten 1993; ‘t Hart et al. 1996; Rahman et al. 2001; Rajasimha et al. 2008). Mouse models have also been utilized (Cree et al. 2008; Fan et al. 2008; Freyer et al. 2012; Ross et al. 2013), but to date, there have been only a few studies of normal patterns of heteroplasmy transmission in humans (Sekiguchi et al. 2003; Goto et al. 2011; Sondheimer et al. 2011; Guo et al. 2013; Rebolledo-Jaramillo et al. 2014), including studies of oocytes and placenta (Marchington et al. 1997, 2002; Jacobs et al. 2007), and several questions remain.For example, although it is clear that a bottleneck occurs during the transmission of mtDNA genomes from mothers to offspring, the size of the bottleneck remains a contentious issue. Previous estimates of the effective number of transmitted mtDNA genomes range widely, from eight to 200 (Brown et al. 2001; Guo et al. 2013; Rebolledo-Jaramillo et al. 2014). However, all previous studies have assumed a constant size for the bottleneck across individuals; the effect of allowing the bottleneck size to vary among individuals has not been investigated. Moreover, it has been suggested that mtDNA genomes may not behave as independent entities but instead are organized into discrete units called “nucleoids,” each of which contains 5–10 mtDNA genomes (Jacobs et al. 2000; Cao et al. 2007; Khrapko 2008), although recently it has been suggested that the number may be much smaller, on the order of one mtDNA genome per nucleoid (Kukat et al. 2011). Each nucleoid is thought to be homoplasmic for mtDNA genome sequences; thus, mtDNA heteroplasmy at the cellular level would reflect nucleoids that are homoplasmic for different sequence variants. Nucleoid structures within cells have been studied microscopically and biochemically (Bogenhagen 2012), and nucleoid-based models have been found to provide a better fit to the segregation of heteroplasmic mtDNA genomes in cell lines than do simple bottleneck models in some studies (Cao et al. 2007; Khrapko 2008), but not in others (Cree et al. 2008). However, to date, nucleoid-based models have not been investigated in the transmission of mtDNA heteroplasmy from mothers to offspring.Another issue is the degree to which negative (or purifying) selection may act on deleterious variants during the transmission of mtDNA heteroplasmy. There are conflicting results and views as to whether changes in the frequency of a heteroplasmic mutation from mother to offspring are governed solely by genetic drift, or whether there is an additional role for negative (purifying) selection (Jenuth et al. 1997; Durham et al. 2006; Stewart et al. 2008a,b; Wonnapinij et al. 2008; Wallace and Chalkia 2013; Rebolledo-Jaramillo et al. 2014). Negative selection during heteroplasmy transmission, as evidenced by a decrease in the frequency of presumably deleterious heteroplasmic variants in offspring compared to mothers, must operate on the female germ line and/or early in development after fertilization, and hence differs from negative selection operating on homoplasmic variants that reduce viability or fertility (Holt et al. 2014). The opportunities for, and extent of, such negative selection during heteroplasmy transmission in humans remain largely unknown.Here, we utilize the Genomes of the Netherlands (GoNL) project (Boomsma et al. 2014; Genome of the Netherlands Consortium 2014), consisting of whole-genome sequence data from blood samples from 250 families, to carry out the largest study to date (to our knowledge) of the dynamics of heteroplasmy transmission across the entire mtDNA genome. We utilize the data on changes in minor allele frequency (MAF) from mothers to offspring at heteroplasmic sites to compare different models for the inheritance of mtDNA genomes, and we analyze the data for evidence of negative selection during heteroplasmy transmission.     

Temporal discrimination of two passive movements in humans: a new psychophysical approach to assessing kinaesthesia

Percutaneous electrical stimulation of the motor point of the first dorsal interosseous muscle (FDI) was used to produce a non-painful contraction of the FDI muscle that caused index finger abduction movement but no radiating cutaneous paraesthesias or sharp sensations localized to joints. Pairs of stimuli separated by different time intervals were given and subjects were asked to report whether they perceived a single or a double index finger abduction movement. The threshold value was the shortest interval for which the subjects reported two separate index finger abduction movements. Temporal discrimination movement thresholds (TDMT) were measured for both right and left hand. To assess the possible role of muscle and cutaneous afferents in temporal discrimination, we investigated the effects of high-frequency (20 Hz) electrical stimulation of the right ulnar and radial nerves on TDMT. In humans, muscle afferents from FDI are supplied by the ulnar nerve whereas the cutaneous territory overlying the muscle and joint is supplied by the radial and median nerves. Threshold values were not significantly different for right (75.1 ms) and left (75.6 ms) hands. During ulnar and to a lesser extent during radial nerve stimulation, TDMT values were significantly increased (119.2 and 93.5 ms, respectively) compared with baseline conditions (78.0 ms) whereas no changes were observed during median nerve stimulation (80.5 ms). These results suggest that muscle, and in part cutaneous, afferents contribute to temporal discrimination of a dual movement. The technique may provide a useful way of measuring temporal discrimination of kinaesthetic inputs in humans.     

The salivary microbiota as a diagnostic indicator of oral cancer: A descriptive,non-randomized study of cancer-free and oral squamous cell carcinoma subjects

Background  

The purpose of the present investigation was to determine if the salivary counts of 40 common oral bacteria in subjects with an oral squamous cell carcinoma (OSCC) lesion would differ from those found in cancer-free (OSCC-free) controls.     

A genomic analysis of chicken cytokines and chemokines.

As most mechanisms of adaptive immunity evolved during the divergence of vertebrates, the immune systems of extant vertebrates represent different successful variations on the themes initiated in their earliest common ancestors. The genes involved in elaborating these mechanisms have been subject to exceptional selective pressures in an arms race with highly adaptable pathogens, resulting in highly divergent sequences of orthologous genes and the gain and loss of members of gene families as different species find different solutions to the challenge of infection. Consequently, it has been difficult to transfer to the chicken detailed knowledge of the molecular mechanisms of the mammalian immune system and, thus, to enhance the already significant contribution of chickens toward understanding the evolution of immunity. The availability of the chicken genome sequence provides the opportunity to resolve outstanding questions concerning which molecular components of the immune system are shared between mammals and birds and which represent their unique evolutionary solutions. We have integrated genome data with existing knowledge to make a new comparative census of members of cytokine and chemokine gene families, distinguishing the core set of molecules likely to be common to all higher vertebrates from those particular to these 300 million-year-old lineages. Some differences can be explained by the different architectures of the mammalian and avian immune systems. Chickens lack lymph nodes and also the genes for the lymphotoxins and lymphotoxin receptors. The lack of functional eosinophils correlates with the absence of the eotaxin genes and our previously reported observation that interleukin- 5 (IL-5) is a pseudogene. To summarize, in the chicken genome, we can identify the genes for 23 ILs, 8 type I interferons (IFNs), IFN-gamma, 1 colony-stimulating factor (GM-CSF), 2 of the 3 known transforming growth factors (TGFs), 24 chemokines (1 XCL, 14 CCL, 8 CXCL, and 1 CX3CL), and 10 tumor necrosis factor superfamily (TNFSF) members. Receptor genes present in the genome suggest the likely presence of 2 other ILs, 1 other CSF, and 2 other TNFSF members.     

Discordant and heterogeneous clinically relevant genomic alterations in circulating tumor cells vs plasma DNA from men with metastatic castration resistant prostate cancer

Circulating tumor cell (CTC) and cell‐free (cf) DNA‐based genomic alterations are increasingly being used for clinical decision‐making in oncology. However, the concordance and discordance between paired CTC and cfDNA genomic profiles remain largely unknown. We performed comparative genomic hybridization (CGH) on CTCs and cfDNA, and low‐pass whole genome sequencing (lpWGS) on cfDNA to characterize genomic alterations (CNA) and tumor content in two independent prospective studies of 93 men with mCRPC treated with enzalutamide/abiraterone, or radium‐223. Comprehensive analysis of 69 patient CTCs and 72 cfDNA samples from 93 men with mCRPC, including 64 paired samples, identified common concordant gains in FOXA1, AR, and MYC, and losses in BRCA1, PTEN, and RB1 between CTCs and cfDNA. Concordant PTEN loss and discordant BRCA2 gain were associated with significantly worse outcomes in Epic AR‐V7 negative men with mCRPC treated with abiraterone/enzalutamide. We identified and externally validated CTC‐specific genomic alternations that were discordant in paired cfDNA, even in samples with high tumor content. These CTC/cfDNA‐discordant regions included key genomic regulators of lineage plasticity, osteomimicry, and cellular differentiation, including MYCN gain in CTCs (31%) that was rarely detected in cfDNA. CTC MYCN gain was associated with poor clinical outcomes in AR‐V7 negative men and small cell transformation. In conclusion, we demonstrated concordance of multiple genomic alterations across CTC and cfDNA platforms; however, some genomic alterations displayed substantial discordance between CTC DNA and cfDNA despite the use of identical copy number analysis methods, suggesting tumor heterogeneity and divergent evolution associated with poor clinical outcomes.     

Memory for fingertip forces: passive hand muscle vibration interferes with predictive grip force scaling

When subjects repetitively lift an object, the grip force they select is influenced by the mechanical object properties of the preceding lift. Similar effects on grip force scaling are observed whether the subsequent lift is performed with the same hand or the hand contralateral to the preceding lift. Here we demonstrate that passive vibration of the hand muscles involved in the generation of grip force in the interval between two blocks of lifting trials interferes with predictive grip force scaling. Following ten trials in which subjects lifted an object with constant mechanical properties with the dominant hand, muscle vibration was given to the first interosseus and adductor pollicis muscles of the dominant hand during a 10-min rest period. Compared with the last lift preceding vibration, peak rates of grip force increase and peak grip forces were scaled too high during the first lift following vibration whether the lift was made with the dominant or non-dominant hand. Subjects scaled grip force accurately to the object properties within three lifts following vibration. If subjects rested for 10 min after the first ten trials and received no vibration, then there was no significant difference in the peak grip force or its rate of increase between the last lift preceding rest and the first lift following it. We suggest that vibration impairs the memory processes responsible for predictive grip force scaling. Our data are consistent with the recent suggestion that these memory processes are neither specific for a certain motor action nor do they reflect internal representations of mechanical object properties.     

The long-term effectiveness of generic adult fixed-dose combination antiretroviral therapy for HIV-infected Ugandan children

Background

Access to pediatric antiretroviral formulations is increasing in resource-limited countries, however adult FDCs are still commonly used by antiretroviral therapy (ART) programs.

Objective

To describe long-term effectiveness of using adult FDC of d4T+3TC+NVP (Triomune) in children for HIV treatment.

Methods

Clinical, immunologic, and virologic outcomes of HIV-infected ART-naïve children aged six months to 12 years, were evaluated up to 96 weeks post-ART initiation.

Results

From March 2004 to June 2006, 104 children were followed with a median age of 5.4 years, median CD4 cell percent and HIV-1 RNA were 11.0% (IQR 6.7–13.9) and 348,846copies/mL (IQR 160,941–681,313) respectively at baseline. Using Kaplan-Meir estimates, 75% of children had undetectable viral loads (<400copies/mL) at 96weeks of ART. Children with a baseline CD4 cell percent >15% were 3 times more likely to achieve viral load <400copies/mL than those with baseline CD4 cell percent <5% after adjusting for baseline age {aHR = 3.03 (1.10–8.32), p=0.03}; no difference was found among those with CD4 cell percent >5–14.9% and <5%.

Conclusion

Treatment with generic adult FDC for HIV-infected Ugandan children led to sustained clinical, immunologic and virologic response during 96 weeks of ART. Early initiation of ART is key to achieving virological success.