Human menopause is an unsolved evolutionary puzzle, and relationships among the factors that produced it remain understood poorly. Classic theory, involving a one-sex  model of human demography, suggests that genes imparting deleterious effects on post-reproductive survival will accumulate. Thus, a ‘death barrier’ should emerge beyond the maximum age for female reproduction. Under this scenario, few women would experience menopause  because few would survive much longer than they reproduced. However, no death barrier is observed in human populations. Subsequent theoretical research has shown that two-sex models, including male fertility at older ages, avoid the death barrier. Here we use a stochastic, two-sex computational model implemented by computer simulation to show how male mating preference for younger females could lead to the accumulation of mutations deleterious to female fertility and thus produce a menopausal period. Our model requires neither the initial assumption of a decline in older female fertility nor the effects of inclusive fitness through which older, non-reproducing women assist in the reproductive efforts of younger women. Our model helps to explain why such effects, observed in many societies, may be insufficient factors in elucidating the origin of menopause. The origin and evolution of menopause is understood poorly and explanations remain contentious. Virtually ignored among explanations is the effect that mate choice can exert on an evolving population. We designed and used a computational model and computer simulation to show that male mating preference for younger females in humans could have led to the accumulation of mutations deleterious to female fertility and thereby produced menopause. Our model demonstrates for the first time that neither an assumption of pre-existing diminished fertility in older women nor a requirement of benefits derived from older, non-reproducing women assisting younger women in rearing children is necessary to explain the origin of menopause. Evolutionary theories of life history predict that selection should operate against living beyond the age of reproduction Solutions may be grouped into two general explanatory categories ij], connected male and female life histories. The mating preference matrix represented the propensity for a male of age i to form a bond with a female of age j, which might have yielded offspring if both were fertile. Such two-sex models are mathematically much more complex than are one-sex models because of the inherent non-linearity introduced by the mating preference matrix. Tuljapurkar et al. Hamilton and fertility, and we established mutation-selection balance equilibria that were effected by mate choice and affected fertility. We implemented mutant alleles at any among five autosomal loci that imparted nonpleiotropic, deleterious effects . These five loci affected survival probability for individuals in age classes beyond a particular age of onset. Individual survival thus was determined by the number and kind of mortality-causing mutations that had accumulated in the population in which a male or female were a member and, ultimately, had been inherited (details are provided in the section “Model” and tables in the Supporting Information). Eventually, simulations produced mutation-selection balance  in which the introduction of new mutations was balanced by the elimination, through stochastic deaths, of existing mutations. Deaths were compensated by births from pseudo-randomly chosen mating pairs. Mating pairs were formed according to an age-dependent mating preference matrix. Pairs could produce offspring only if both male and female parents were fertile, as determined from age-dependent probabilities in fertility tables. Model AP involved an age indifferent preference in the formation of mating pairs  combined with extended male fertility prevents an early decline in female survival . How did this arise? Some evidence suggests that ancestral human populations were characterized by intrinsic female menopause ijAP, sex-specific, infertility-causing mutations . As a result of these mutations, female fertility declined before female survival, and female menopause arose  to a matrix encoding preference for younger females , this time involving a single locus with age of onset in class 6 . Larger mutation rates produced more-rapid approaches to fixation when selection was reduced . During the mutation-selection balance period (model AP), the equilibrium frequency was higher for higher mutation rates. Increasing the selection strength reduced the equilibrium frequency of mutations  but imparted little effect on the approach to fixation during relaxed selection (YF). Decreasing the population size imparted little effect (The difference between age indifferent preference (model AP) and preference for younger females (model YF) in producing a menopausal effect was robust across population sizes and mutation rates. We next explored the effect of shifting from a matrix encoding age indifferent preference  is assigned to an age class (1 to 18). Each individual is associated with a diploid genotype encoded at a number of loci. Mutations at a specific locus may affect either fertility or survival. Mutations may act in a sex-indifferent  or sex-specific  manner defined by an age of onset (for the locus) and with a fertility table.Initially, individuals are assigned pseudo-randomly to age classes, without mutations. Then, a ‘burn-in’ period allows the age distribution to reach an approximate steady state.Each time interval (representing a 5-year period) consists of determining a survival probability for each individual according to an intrinsic survival table, modified by the number and kind of mortality-affecting mutations carried. A pseudo-random number is used with the computed survival probability to determine if each individual survives and is assigned to the next age class or dies. All individuals in age class 18 die. Deaths are replaced in the population by new births assigned to the first age class.ij] is applied to potential parents (based on their age classes i and j) to determine the probability of pair formation. Parent male and female fertility probabilities are determined using intrinsic fertility tables, modified by the number and kind of mutations carried. A pseudo-random number is used to determine if a birth occurs. If not, the birth simulation process is repeated until potential parents become actual parents.Births are simulated by selecting a potential father pseudo-randomly from the surviving male population and a potential mother pseudo-randomly from the surviving female population. The appropriate ‘mating preference matrix’ [MParents transmit their alleles to offspring according to autosomal Mendelian genetic rules; thus, if a parent is homozygous, then that parent transmits one mutation; if heterozygous, then the mutant allele is transmitted with 50% probability. At this time, the introduction of new mutations into each newborn is simulated using a pseudo-random number and the appropriate mutation rate.Simulations were run on a Power Macintosh G5 personal computer (running OS X 10.5.8), using the C programming language (with the Apple GCC compiler). Computational model code is accessible online in the Supporting Information .Table S1Intrinsic fertility and survival probabilities.(DOC)Click here for additional data file.Table S2AP.Mating preference matrix M(XLS)Click here for additional data file.Table S3YP.Mating preference matrix M(XLS)Click here for additional data file.Table S4Intrinsic fertility (prolonged) and survival.(DOC)Click here for additional data file.Table S5Female-specific, infertility-causing loci.(DOC)Click here for additional data file.Table S6Sex-indifferent, mortality-causing mutations.(DOC)Click here for additional data file.Text S1Computational model code.(TXT)Click here for additional data file.


Vacuoles of plants fulfill various biologically important functions, like turgor generation and maintenance, detoxification, solute sequestration, or protein storage. Different types of plant vacuoles (lytic versus protein storage) are characterized by different functional properties apparently caused by a different composition/abundance and regulation of transport proteins in the surrounding membrane, the tonoplast. Proteome analyses allow the identification of vacuolar proteins and provide an informative basis for assigning observed transport processes to specific carriers or channels. This review summarizes techniques required for vacuolar proteome analyses, like e.g., isolation of the large central vacuole or tonoplast membrane purification. Moreover, an overview about diverse published vacuolar proteome studies is provided. It becomes evident that qualitative proteomes from different plant species represent just the tip of the iceberg. During the past few years, mass spectrometry achieved immense improvement concerning its accuracy, sensitivity, and application. As a consequence, modern tonoplast proteome approaches are suited for detecting alterations in membrane protein abundance in response to changing environmental/physiological conditions and help to clarify the regulation of tonoplast transport processes. Plant vacuoles can be distinguished by their size, specific metabolic properties, state of maturation, etc. Moreover, their number and physiological role can undergo dramatic changes during cell cycle and development   .In 2007, a very detailed description of intact vacuole isolation from Vacuole isolation from protoplasts might be considered as problematic because cell wall lysis takes about 2–4 h and hence might represent a stress situation for the cells and generated protoplasts. Therefore, it is imaginable that some analyses are affected by this putative stress situation. Particularly, comparative studies, including quantitative proteome investigations of vacuoles isolated from plants after application of abiotic or biotic stress factors, have to be critically evaluated. Application of suitable controls performed in parallel helps to identify “artifacts” caused by the isolation method. Metabolite measurements of vacuoles isolated from cold acclimated and control plants demonstrated a significant increase in glucose and fructose concentrations solely in those vacuoles that were isolated from the plants incubated at 4°C  for 8 days. Cell wall digestion and generation of protoplasts was conducted as described in previous studies  but also the number of detected peptides (of a single protein) as well as protein coverage have to be considered.Arabidopsis genome encodes 28 proteins proven or at least predicted to represent V-type ATPase subunits, including different subunit isoforms , almost all qualitative vacuolar proteome analyses contained well-known plastidial and mitochondrial proteins, like the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase, chlorophyll binding proteins of the light harvesting complex, and the mitochondrial ATP-synthase or the ADP/ATP carrier  is part of the salt-overlay-sensitive (SOS) pathway and was shown to activate the plasma membrane resident Na+/H+ antiporter SOS1  . At least one phosphopeptide was identified in 130 phosphorylated proteins. 58 phosphorylated proteins contained one or more transmembrane domains and hence were classified as membrane proteins. Twenty-eight of the 58 phosphorylated membrane proteins represent transport proteins specific for anions, sugars, potassium, sodium, or oligopeptides as well as ABC transporters   protein kinases were identified in the tonoplast phosphoproteome proteomic analysis of the plasma membrane and the tonoplast of Oryza sativa was published  suggests that this type of regulation is at least partially conserved between monocotyledones and dicotyledones. However, it is also imaginable that rice and maybe other monocot groups exhibit specific phosphorylation of proteins because several phosphopeptides were exclusively detected in rice  or TiO2 affinity chromatography. Sixty-five identified phosphopeptides covered 27 tonoplast proteins of known biochemical function, such as two vacuolar proton pumps, aquaporins, calcium exchangers, Na+/H+ antiporters, and transport proteins for potassium, malate, sulfate, and sugar  generally allows faster regulation. Therefore, the identification of phosphorylated tonoplast proteins is consistent with the physiologically required flexibility of vacuolar transport regulation. Continuative  investigations will provide deeper insights into post-translational regulation of vacuolar transport and might discover further principles  controlling vacuolar function.AtTMT1-loop and to stimulate sugar uptake into the vacuole  provided the basis for the identification of the interacting RAF-type MAPKKK-like kinase. This kinase was shown to phosphorylate at least the So far only few quantitative tonoplast/vacuolar proteome studies have been carried out.isobaric tags for relative and absolute quantitation, iTRAQ), or specific amino acid residues can be labeled with stable isotopes . Moreover, it is also possible to label proteins in living organisms by feeding stable isotopes . This labeling procedure is not applicable for plants grown on soil and requires hydroponic plant cultivation or plant cell cultures. It is also possible to conduct a label-free quantification. An important advantage of a label-free strategy is the generally higher sensitivity (higher number of identified proteins). However, experimental replications and comparably demanding and extensive statistical analyses are necessary.Four different approaches are generally applied to distinguish between proteins from the different samples and to allow comparison of protein abundances  resulted in increased abundance of six proteins, the vacuolar H+ pyrophosphatase (gi|11527561), a TIP1-like protein (gi|520936), a putative natural resistance-associated macrophage protein (NRAMP) (gi|28865876), the vacuolar cation/proton exchanger CAX1a (gi|73917674), a multidrug-resistance(MRP)-like ABC transporter (gi|27368887), and an uncharacterized membrane protein (gi|115452029). In contrast to low Cd2+ concentrations, high Cd2+ concentrations (200 μM) apparently resulted in a more specific response because solely the abundance of the MRP-like protein was increased  photosynthesis  and one peripheral subunit (VHA-E1) of the V-type ATPase, exhibited higher abundance. Enzyme activity measurements were conducted to substantiate the involvement of V-type ATPase in cold acclimation. In fact, ATPase activity in tonoplast vesicles was considerably enhanced (at least fourfold) due to cold exposure. Therefore, the higher abundance of V-type ATPase subunits apparently results in a higher enzymatic activity. Cold acclimation resulted in higher abundance of nine solute transporters, including well-characterized tonoplast carriers, like the dicarboxylate transporter AtTDT, the high affinity nitrate transporter AtNRT2.7, and the zinc/H+ antiporter AtMTP1/ZAT1 (Schulze et al., AtTDT (Emmerlich et al., AtTDT.A quite recent study focused on the role of the large central vacuole in adaptation to cold. Decreasing environmental temperatures initiate a process called cold acclimation (Schulze et al., t et al. . Four biAtTMT1 and AtTMT2 (Wormit et al., AtTMT proteins revealed that cold acclimation stimulates phosphorylation of both, AtTMT1 and AtTMT2. Therefore, increased vacuolar accumulation of monosaccharides might result from higher activity of the TMT proteins caused by phosphorylation. Proteome data in combination with physiological studies for the first time clearly demonstrated that the vacuole is involved in the process of cold acclimation (Schulze et al., Determination of soluble sugar contents revealed that cold acclimation results in several-fold higher concentrations of sucrose, glucose and fructose, and that accumulation of the monosaccharides largely occurs in the vacuole. Interestingly, abundance of the main vacuolar monosaccharide transporters Arabidopsis t-DNA-insertion lines lacking certain protein kinases might be a promising approach for the identification of signal pathways controlling vacuolar protein phosphorylation. A further challenge will be the large-scale investigation of the protein composition of non-lytic vacuoles, like protein storage vacuoles or pre-vacuolar vesicles. Comparison of the respective results might help to define differences between different types of vacuoles.Proteome studies considerably broadened the knowledge about the large central vacuole of plants. Qualitative proteome analyses identified various vacuolar and particularly, tonoplast proteins with increasing accuracy. Based on these important findings plant scientists managed to clarify the biochemical function and physiological relevance of several vacuolar proteins. Phosphoproteome data and physiological studies demonstrated that vacuolar proteins are modified by phosphorylation and that post-translational modification might represent an important mechanism allowing fast adaptation of vacuolar functions in accordance to the cellular demands. Comparative proteome studies elucidated the regulatory impact of abiotic stresses on vacuolar protein abundance. Sophisticated comparative proteome studies, might help to identify specific protein modifications induced by changing conditions. Investigation of tonoplast protein phosphorylation in The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Over the past several years meditation practice has gained increasing attention as a non-pharmacological intervention to provide health related benefits, from promoting general wellness to alleviating the symptoms of a variety of medical conditions. However, the effects of meditation training on brain activity still need to be fully characterized. Sleep provides a unique approach to explore the meditation-related plastic changes in brain function. In this study we performed sleep high-density electroencephalographic (hdEEG) recordings in long-term meditators (LTM) of Buddhist meditation practices (approximately 8700 mean hours of life practice) and meditation naive individuals. We found that LTM had increased parietal-occipital EEG gamma power during NREM sleep. This increase was specific for the gamma range (25–40 Hz), was not related to the level of spontaneous arousal during NREM and was positively correlated with the length of lifetime daily meditation practice. Altogether, these findings indicate that meditation practice produces measurable changes in spontaneous brain activity, and suggest that EEG gamma activity during sleep represents a sensitive measure of the long-lasting, plastic effects of meditative training on brain function. Meditation can be conceptualized as a set of regulatory and self-inquiry mental training regimes cultivated for various ends, including the training of well-being and psychological health An important question raised by these studies is whether enhanced EEG activity is state dependent (e.g.– occurring only during meditation and immediately after it), or trait dependent  The recent availability of high-density electroencephalography (hdEEG) has greatly enhanced the spatial resolution of standard EEG recordings, thus allowing for better characterization of local plastic changes Here we performed whole night sleep hdEEG (256-channels) in experienced meditators (LTM) and meditation naive individuals matched for age and sex. Even if this is a correlative and not a causal approach, it paves the way for studies on long term plastic effects of meditation on brain activity in healthy humans.Twenty-nine right-handed long-term meditators  and a group of twenty-nine meditation naive subjects matched for age and sex were recruited. LTM had a history of daily meditation practice of at least 3 years and had participated in at least 3 one-week intensive retreats. Mean duration of meditation training was 15.6 years . Naive subjects had no previous experience with meditation. All LTM participants were proficient in meditation practices, as taught within the framework of either Theravada or Tibetan Buddhisms. These practices included two attention-based meditations, which we referred to as open monitoring (OM) and focused attention (FA), as well as one compassion/loving kindness meditation referred to as metta meditation All subjects underwent in-laboratory hdEEG polysomnography (PSG) that utilized 256 channel hdEEG , as well as standard sleep monitoring leads, including electrooculogram, sub-mental electromyogram, electrocardiogram, bilateral tibial electromyogram, respiratory inductance plethysmography, pulse oximetry, and a position sensor. Participants arrived at the laboratory between 4:00 and 5:00 p.m. for set-up that took approximately two hours. Baseline EEG and an attention and a fear conditioning task were performed. After 9pm the participants were allowed to sleep undisturbed in the laboratory beginning within one hour of their usual bedtime. Additional measures were collected the next day. In this report we will focus only on the EEG data during sleep.The socioeconomic status (SES) measure  meditation practice per week, including time spent on meditation retreats , a sub-mental electromyogram and an electrooculogram, was performed by a registered polysomnographic technologist in 30-second epochs according to standard criteria Sleep cycles were defined according to the modified criteria Differences in clinical as well as sleep architecture variables were examined using 2-tailed, unpaired t-tests. Topographical analysis was computed after spatially normalizing each subject's topography  within the frequency bands of interest as a means to reduce between-subject variance. Group differences in topographical NREM and REM sleep hdEEG power were assessed with statistical non-parametric mapping (SnPM) using a suprathreshold cluster test to identify significant groups of electrodes after accounting for the multiple comparisons due to the numerous electrodes The magnitude of NREM sleep EEG gamma power differences between LTM and meditation naive individuals was assessed with the Cohen's d, a measure of the effect size The LTM and the meditation naive groups were matched for age and sex see . AdditioWe first checked for global, frequency non-specific differences, by comparing the absolute average power across the spectrum between the groups. This analysis revealed no difference between LTM and meditation naive individuals (p = 0.094). We then focused on local, frequency specific effects. Whole night NREM sleep EEG power was topographically compared between LTM and meditation naives by spatially normalizing each subject's map to the average power in six frequency ranges . A frequency specific increase in EEG gamma power was found in LTM. Whereas NREM sleep gamma power in both LTM and meditation naives was strongest in frontal/prefrontal areas, and weakest in the temporal regions bilaterally , top rowWe found no gamma power difference in either the electromyographic (EMG) or electrooculographic (EOG) derivations between the two groups (p = 0.589 and p = 0.493 respectively) , suggestTo further characterize the gamma increase at the individual subject level, we calculated the average NREM gamma power of the parietal-occipital cluster for each subject. The average cluster difference in gamma power between groups was 35%  and the To assess whether the gamma increase of LTM was specific for NREM sleep, topographic EEG analysis of REM sleep was also performed. As previous literature has suggested functional differences between tonic and phasic REM, we separated these two REM sleep patterns in our analysis By performing sleep hdEEG (256 channels) recordings in this study we found that LTM had increased parietal-occipital EEG gamma power during NREM sleep compared to meditation naives. This increase was specific for the gamma frequency range, was not found during REM sleep, and was positively correlated with the length of daily meditation practice.The LTMs recorded in this study were experienced in Focused Attention (FA), Open Monitoring (OM), as well as loving kindness/compassion meditations. An increase in gamma activity was recently demonstrated in expert Buddhist practitioners (>10.000 hours of practice) during a style of meditation which contains features of both loving kindness/compassion and OM meditations Here we established that LTM had higher EEG gamma power over an extended period of spontaneous brain activity  compared to meditation naive individuals. This parietal-occipital gamma increase corresponded to a large effect size . These findings strongly suggest that changes in EEG gamma activity related to meditation practice are trait (in addition to state) related. Comparing the activity of meditation experts and novices during an unambiguous resting condition is a challenging task Daily practice and meditation retreat could contribute differently to the neuroplastic changes induced by meditation. For instance, meditation frequency (days per week with meditation practice) has recently been shown to reliably predict both higher mindfulness and psychological well-being In the present study LTM had a significantly reduced total sleep time (TST) and increased wake after sleep onset (WASO) compared to meditation naive individuals. A reduction in TST has been recently reported by another sleep study in LTM Only a handful of studies so far have investigated the sleep EEG activity of meditation experts beyond sleep architecture. One of these studies explored EEG differences in thirteen individuals with at least 2 years of meditation experience during Transcendental meditation , resting wakefulness, drowsiness, and sleep and found a progressive slowing of the main EEG frequency from wakefulness to sleep, with no appreciable change in power between wakefulness and meditation EEG What is the functional meaning of the gamma increase in LTM that was found here during NREM sleep? A large body of evidence from animal and human recordings have suggested that gamma-frequency activity is implicated during wakefulness in plasticity-related processes, including attention, learning, as well as both working and long-term memory Previous studies have reported an increase in fast-frequency activity during sleep in pathological conditions such as schizophrenia, depression and insomnia The increase in NREM sleep parieto-occipital gamma power reported here could also reflect an enhanced activity of the underlying cortical areas in LTM compared to meditation naives. Higher EEG gamma power reflects higher firing rates of the underlying neuronal populations Limitations of the study include a lack of an adaptation night, which could account for the truncated sleep time (< 7 hrs) in all participants, but it is unlikely to explain the observed group difference in the sleep EEG. Future work will also need to address some of the questions left unanswered in the present study. For example, the relationship found here between higher EEG gamma activity and longer meditation daily practice suggests that gamma power is a good correlate of meditation training. This correlation should be confirmed in longitudinal studies performing EEG recordings in meditation naive individuals before and after meditation training, ideally using only one style of meditation practice. It will also be important to investigate whether the observed gamma increase may be affected by pre-existing "baseline" gamma activity differences between groups , as previously suggested Figure S1REM tonic as well as phasic gamma power did not differ between LTM and meditation naives. Topographic color plots showed maximal REM tonic as well as phasic gamma power in frontal/prefrontal regions in both groups. Compared to meditation naives, LTM had a slightly higher power in the same parieto-occipital region significantly more active during NREM sleep, which however failed to reach significance in both tonic and phasic REM . Only the tonic REM topographies are shown.(TIF)Click here for additional data file.


Objectives: Following radical oncological resection, full-thickness upper central trunk defects present a significant challenge. Common reconstructive options include pedicled flaps, such as pectoralis major, rectus abdominis, and latissimus dorsi. In complex cases, free tissue transfer may be required. Reverse abdominoplasty, although initially described for cosmetic body contouring, can be used to reconstruct upper central trunk defects following radical tumour ablation. We present 4 such applications in the management of advanced or recurrent malignancies and review the relative indications for this approach. Methods: Four consecutive cases (2004-2010) were reviewed with respect to indication, operative procedure, and complications. Results: There were no cases of complete flap loss. One patient underwent revision for marginal flap necrosis while another developed local recurrence, requiring re-excision and reconstruction with flap advancement. Conclusions: Where pedicled flaps are unavailable or insufficient, adjacent abdominal tissue can be recruited into chest wall defects, avoiding microsurgical free tissue transfer. The authors feel that the reverse abdominoplasty is currently underused in this context and offers an excellent alternative in complex cases where other reconstructive options are unavailable, or where comorbidities preclude free-tissue transfer. The technique is versatile, simple to perform and affords an acceptable cosmetic outcome, yet is not widely reported in the literature. It has particular merit in cases with a high chance of disease recurrence, in the management of recurrent breast cancer, and in patients with multiple comorbidities. The reverse abdominoplasty should therefore be considered when evaluating patients for oncological trunk reconstruction. Central trunk defects following radical ablation of malignancy pose a reconstructive challenge. This is primarily due to the limited availability of local flaps, their reach restrictions, and the uncertainty about achieving clear excision margins. The situation may be further complicated by disease recurrence and the adverse effects of previous radiotherapy on surrounding tissues.-5-7-The reverse abdominoplasty flap, first described by Rebello and Franco,2From 2004 to 2010, 4 reverse abdominoplasty procedures were performed by the senior author for reconstruction of full-thickness upper central trunk defects following oncological resection. The patients, all female, had a mean age of 50 years  at the time of the initial reverse abdominoplasty procedure. Three cases were performed as elective procedures whereas, in one case, paraneoplastic disease necessitated “emergency” mastectomy and reconstruction. Two patients had received local radiotherapy prior to the procedure.The average period of follow-up from operation to the time of writing was 47 months. There were no cases of complete flap loss. One patient underwent surgical revision for marginal flap necrosis while another developed local recurrence, which required re-excision and further reconstruction.top left). The abdominoplasty flap is raised below Scarpa's fascia, just above the rectus sheath (bottom left). The dissection continues beyond the umbilicus to the pubic area and the groin creases on each side. The umbilical stalk is usually divided to allow the umbilicus to “float.” Dissection is aided by the use of a deep or long-lighted retractor or a headlight. Meticulous hemostasis is achieved and 2 suction drains are inserted. The suction drains are placed deep to the advanced flap, carefully avoiding the superficial inferior epigastric vessels. The caudal abdominal tissue is then advanced cranially into the chest wall defect (bottom right). The flap is inset using 2/0 PDS sutures from the Scarpa's layer to the superficial fascial system of the chest. Skin closure is performed in layers with 3/0 Monocryl deep dermal, running Monocryl subcuticular followed by Steri-strips  applied across the wound.Preoperative assessment of abdominal wall laxity is carried out with the patient in both the standing and the recumbent positions. The inferior resection margin of the tumor ablation constitutes the superior border of the abdominoplasty flap and must be adequate and not compromised by concerns to preserve tissue for the reconstruction. The markings may be subsequently revised during surgery according to final size and configuration of the defect present. An inframammary “gull-wing” incision is made, centered on the midline . She remains disease-free and has declined delayed breast reconstruction.The postoperative course was complicated by a small area of fat necrosis in the distal end of the flap, which necessitated debridement and direct (primary) closure. The final postoperative appearances are satisfactory , bottom.A 48-year-old woman presented with a progressively enlarging lump in her upper central abdomen. Her surgical history included an uncomplicated laparoscopic cholecystectomy for gallstones 5 years earlier. Computed tomographic images showed that the mass extended posteriorly to the anterior wall of the stomach and a core biopsy confirmed an adenocarcinoma; the pathological diagnosis was a port-site metastasis originating from a gallbladder tumor. During the next 3 weeks, the tumor enlarged rapidly reaching a palpable 15 × 12 cm epigastric mass with clinically detectable diastasis recti.A full-thickness resection of the tumor in the anterior abdominal wall was carried out under frozen section guidance. The lesion was entirely contained within the falciform ligament, but it was necessary to include segments of rectus abdominis muscle in the resection. The resulting defect was reconstructed prosthetically in 2 layers comprising a posterior DynaMesh  and an anterior Prolene mesh . Skin and soft tissue coverage was achieved with a reverse abdominoplasty advancement flap . There wA 56-year-old woman underwent breast conserving surgery for primary invasive breast cancer, followed by adjuvant breast radiotherapy. Nine years later, she was diagnosed with radiation-induced angiosarcoma of the same breast, for which she underwent bilateral mastectomy. She remained disease free for the next 4 years, when she developed recurrent angiosarcoma of her right chest wall. The recurrence was resected and the chest wall reconstructed using a right latissimus dorsi myocutaneous flap. An involved superior margin necessitated re-excision and direct closure.left). This was resected with a 3-cm margin down to deep fascia, and the defect was reconstructed with a reverse abdominoplasty and full-thickness skin graft (right).A few months later, she manifested a further local recurrence in the upper abdomen/lower chest wall, which proved resistant to a trial of chemotherapy , left. Tin graft , right.top). This necessitated further resection with at least 3-cm margin, including the xiphisternum, perichondria of the lower 3 ribs and the upper parts of the rectus muscles. There was no peritoneal involvement and the central abdominal wall reconstruction was performed using Prolene mesh  and a left latissimus dorsi myocutaneous flap in combination with readvancement of the reverse abdominoplasty flap. Healing was uneventful (bottom) and she remained disease free for the next 4 years.She remained disease free for the next 7 months, before re-presenting with rapid disease progression , top. Theventful , bottom A 32-year-old woman presented to the emergency department with a short history of proximal muscle weakness, a bulbar palsy, and a recently enlarging right breast mass. A computed tomographic scan demonstrated an 8 × 8 cm mass in the medial right breast suggestive of malignancy, with right axillary lymphadenopathy but no evidence of distant metastases. Soft tissue biopsies confirmed a paraneoplastic acute necrotizing myopathy and dermatomyositis.Following a diagnosis of invasive breast cancer, she underwent an urgent right mastectomy and axillary lymph node dissection. This required radical breast skin excision, including the nipple-areolar complex but preserving lateral breast skin. The tumor resection included some pectoralis major muscle for clearance. The resultant defect was closed using a reverse abdominoplasty advancement flap. Histological examination showed a grade III metaplastic carcinoma of the breast with squamous differentiation and extensive lymph node spread.Postoperatively, she required continued intubation and admission to the intensive care unit. She received intravenous steroids and was discharged home 5 weeks after admission.An outpatient review 6 weeks following discharge from hospital revealed a 2 × 8 cm area of dehiscence of the lateral mastectomy skin flap, with an 8 × 5 cm cavity deep to this. This has been treated conservatively with negative pressure wound therapy, although there has been a delay in the commencement of adjuvant treatment due to seroma formation and slow healing.Full-thickness upper central trunk defects following oncological resection present a reconstructive challenge. Local flaps may be unavailable due to involvement of the underlying disease process, whereas flap viability may be compromised by previous or planned chest wall radiotherapy. The potentially recurrent nature of the underlying disease may necessitate wide excision margins followed by multiple ablative and reconstructive procedures. Careful planning is therefore necessary when considering potential and optimal reconstructive options—this may involve local, pedicled or free-flap transfer.-Classification systems9More complex reconstructions, often when local tissues have been severely damaged or the pedicled flaps have insufficient size or reach, may require free tissue transfer.-6,15-18 A search of the English-language literature using PubMed reveals only 2 citations since 1984 describing the technique for reconstruction following oncological resection.7,8The reverse abdominoplasty offers an alternative to pedicled or free flap reconstruction by recruiting adjacent abdominal tissue into the defect. First described in English by Baroudi et al2Although we do not advocate the reverse abdominoplasty as a first-line choice for oncological trunk reconstruction in all cases, we feel that it is currently underused in this context and offers an excellent alternative in complex cases where other reconstructive options are unavailable, or where comorbidities preclude free-tissue transfer.The reverse abdominoplasty has many advantages when used for upper trunk reconstruction . It can The reverse abdominoplasty has particular merit in cases of locally advanced disease where the risk of recurrence is high, or in recurrent breast cancer. Where further resection is necessary, it is much simpler to remove part of the reverse abdominoplasty flap and advance the tissues than to undertake resection of a pedicled or free flap, as is commonly performed. If there is insufficient tissue, the reverse abdominoplasty can be combined easily with other flaps, and the option of free tissue transfer remains available as a last resort. In the case of recurrent breast cancer, the use of the reverse abdominoplasty spares certain potential reconstructive options, such as a pedicled latissimus dorsi flap, which can be considered for secondary reconstruction after a suitable disease-free interval. However, given the advanced and recurrent nature of the cases we describe, none of our patients sought consultation for secondary breast reconstruction.Using this relatively simple technique, we were able to avoid microsurgical free tissue transfer despite the complex nature of primary and recurrent disease described in our case series. This was possible partly because the patients were parous women, who thus had adequate lower abdominal laxity, which could be mobilized to enable abdominoplasty flap advancement superiorly.Following radical oncological resection, full-thickness upper central trunk defects pose a reconstructive challenge. The authors believe that in selected patients, successful reconstruction of central trunk defects following radical tumor ablation can be achieved using the reverse abdominoplasty, without resorting to microsurgical free tissue transfer. We therefore advocate consideration of this technique when evaluating patients for upper central trunk defect reconstruction.


Malignant melanoma represents the third most common cause for cerebral metastases after breast and lung cancer. Central nervous system (CNS) metastases occur in 10 to 40% of patients with melanoma. Most of the symptoms of CNS melanoma metastases are unspecific and depend on localization of the lesion. All patients with new neurological signs and a previous primary melanoma lesion must be investigated. Although primary diagnosis may rely on computed tomography scan, magnetic resonance images are usually used in order to study more precisely the characteristics of the lesions in and to embase the surgical plan. Other possible complementary exams are: positron emission tomography, iofetamine cintilography, immunohistochemistry of liquor, monoclonal antibody immunocytology, optical coherence tomography, and transcriptase-polymerase chain reaction. Treatment procedures are indicated based on patient clinical status, presence of unique or multiple lesions, and family agreement. Often surgery, radiosurgery, whole brain radiotherapy, and chemotherapy are combined in order to obtain longer remissions and optimal symptom relieve. Corticoids may be also useful in those cases that present with remarkable peritumoral edema and important mass effect. Despite of the advance in therapeutic options, prognosis for patients with melanoma brain metastases remains poor with a median survival time of six months after diagnosis. Metastatic spread of tumor cells detached from melanoma into the central nervous system (CNS) occurs haematogenically since lymphatic drainage is absent in the brain . The bloThe cells from brain metastases show a slower growth rate and exhibit lower metastatic potential than cells from visceral metastases, indicating that brain metastases do not necessarily represent the end stage in the metastatical cascade. Rather, brain metastases are likely to originate from a unique subpopulation of cells within the primary neoplasm . Some authors found an association between the size of the cerebral metastatic lesion from malignant melanoma and clinical parameters characteristic of tumor behavior. They classified the metastases from melanomas to their size: smaller than 1 cm (group A), between 1 and 4 cm (group B), and bigger than 4 cm (group C), in order to assess the clinical course of the disease and predict the response to treatment. Group B lesions are the most common, independent of the site of the primary tumor, except for patients with rectal melanoma. Group C metastases are the least common and are usually solitary. Asymptomatic patients usually have group A metastases, whereas those with nonspecific complaints or behavioural changes usually have group B metastases. Solitary lesions usually belong to groups B or C, whereas multiple lesions belong mainly to groups A or B .CNS metastases occur in 10 to 40% of melanoma patients in clinical studies and up to 90% in autopsy studies . The cumulative risk at 5 years for patients with melanoma to develop CNS metastases corresponds to about 7% . Fifty-eSeventy-one percent of the primary lesions are invasive lesions with mean greater than thickness of 3.5 mm. Nevertheless, the studies have showed a significative prevalence of small and well-circumscribed lesions at surgical aspect. There are also observed indolent lesions with a long time of evolution .The case of disseminated carcinomatous cell spreading throughout the brain is called “military metastases” or “carcinomatous encephalitis.” This condition is very rare and correlated with a poor prognosis. Most patients with advanced military metastases will have widespread extracranial disease, but the majority will die from intracerebral spread.Risk factors for central nervous system (CNS) metastases among patients with coetaneous malignant melanoma are: male, head and neck or oral primary lesion, presence of visceral metastases, mainly lung, primary tumor thickness and ulceration of primary lesion. Age and race are not significant factors , 12. One of the many features of the malignant melanoma phenotype, in vitro and in vivo, is the elevated heparanase production and activity, which confers the capacity of degrading the subendothelial matrix produced by endothelial cell monolayer cultures. Supra-additive levels of heparanase activity are found when brain endothelial cells are coincubated with brain-metastatical melanoma cells in equicellular amounts . Murine and human brain-metastatical melanoma cells solubilize sulfated matrix proteoglycans at levels significantly higher than their parental lines. Sulfated matrix proteoglycans are rich in heparan sulfate (HSPGs), with minor amounts of chondroitin and dermatan sulfates. The pattern of HSPG degradation by brain-metastatical melanoma cells differs from that of less metastatical parental cells or cells metastatical to organs other than the brain. Cooperative interactions between heparanases from tumor and endothelial sources are suggested to be a significative mechanism in the invasion process.The brain is a unique microenvironment enclosed by the skull and maintaining a highly regulated vascular transport barrier. To metastasize to the brain, malignant tumor cells must attach to microvessel endothelial cells, invade the blood-brain barrier (BBB), and respond to brain survival and growth factors. Neurotrophins (NTs) are important in brain invasion because they stimulate this process. In brain-metastatic melanoma cells, NTs can promote invasion by enhancing the production of extracellular matrix degradative enzymes such as heparanase, an enzyme capable of locally destroying both the extracellular matrix and the basement membrane of the BBB. Melanoma cell lines exhibiting low ability to form brain metastases express low-affinity neurotrophin receptor p75NTR in relation to their brain-metastatic potentials. Presence of functional TrkC, the putative receptor for the invasion-promoting neurotrophin NT-3, is also expressed in brain-metastatic potential cells.Brain-metastatic melanoma cells can also produce autocrine factors and inhibitors that influence their growth, invasion, and survival in the brain. Synthesis of these factors may influence NTs production by brain cells adjacent to the neoplastic invasion front, such as oligodendrocytes and astrocytes. In brain biopsies, increased amounts of nerve growth factor (NGF) and NT-3 were observed in tumor-adjacent tissues at the invasion front of human melanoma tumors. Astrocytes are supposed to contribute to the brain-metastatic specifity of melanoma cells by producing NTs-regulated heparanase. Trophic, autocrine, and paracrine growth factors may, therefore, determine whether metastatical cells can successfully invade, colonize, and grow in the central nervous system .Integrin alpha(v)beta(3) is a molecule of adhesion to the endothelium, and its expression on the metastatical pattern of human melanoma cells in the central nervous system (CNS) is already studied. Although it is predicted that the adhesion of tumor cells to endothelial cells plays a role in this phenomenon, tumor cell alpha(v)beta(3) integrand expression per se does not explain the difference in metastatic behavior in the CNS. Probably others yet unknown factors must be involved .  Headache is the most common presenting symptom, but brain metastases should be suspected in all melanoma patients with new neurological findings . In patiBrain metastases are clinically diagnosed in the majority of patients with metastatic melanoma. The time from diagnosis of the primary tumor to discovery of disease in the CNS is significantly longer for those who had group A lesions , compared with those who had groups B (metastases between 1 and 1.4 cm) or C lesions (metastases bigger than 4 cm) , 18. In All patient with new neurological signs and a previous primary melanoma lesion must be investigated. CT scan finds solitary lesions in 54.2% and multiple lesions in 45.8% of patients with malignant melanoma cerebral metastasis. Eighty-four percent of the solitary lesions are located in the cerebral hemispheres with 62.5% of these in the frontal region.Seventy-five percent of the CNS metastatic melanoma lesions appear on noncontrast study as increased density; 22% are hypodense, and 3% are isodense. All lesions show contrast enhancement, usually appearing as a homogeneous nodular or ring pattern .MRI is the best diagnostic technique for detecting CNS metastases  Figure . HoweverMost commonly, CNS melanoma metastasis appears in MRI as hyperintense  on T1-wein vivo imaging technique or the time required to obtain histological specimens. Intraoperative identification of brain tumors and tumor margins has been limited by either the resolution of the Optical coherence tomography (OCT) is a new, noncontact, high-speed and high-resolution, real-time, intraoperative imaging technique, capable of resolutions on micrometer scale, which has been used to identify intracortical melanomas. OCT is analogous to ultrasound B-mode imaging, except that reflections of infrared light, rather than sound, are detected. OCT uses inherent tissue contrast, rather than enhancement with dyes, to differentiate tissue types. The compact, fiber-optic-based design is readily integrated with surgical instruments.Two-dimensional images show increased optical backscatter from regions of metastatic melanoma tumors, which are quantitatively used to determine the tumor margin. The images correlate well with the histological findings. Three-dimensional reconstructions reveal regions of tumor penetrating normal cortex and can be re-cut at arbitrary planes. Subsurface cerebral vascular structures can be identified and are then avoided during the surgery. The Literature reports suggest that OCT can effectively differentiate normal cortex from intracortical melanoma based on variations in optical backscatter . High-reDetection and diagnosis of human malignant melanoma by Positron Emission Tomography (PET) using 18F-10B-L-BPA, a specific melanogenesis-seeking compound, has been developed. This resulted in a novel, highly effective methodology for the selective three dimensional imaging of metastatic malignant melanomas, and for accurate determination of 18F-10B concentration in the tumor and surrounding tissue, providing almost all diagnostic information necessary for complete noninvasive radiation dose planning in the treatment of malignant melanoma .The Literature data show that iofetamine (I 123) cintilography can be a good method to differential diagnostic in patients with risk for metastatic melanoma lesions. In patients with metastatic melanoma lesions, cintilography shows increased uptake of iofetamine (I 123). Studies suggest that certain brain tumors such as melanoma are capable of selectively binding iofetamine I 123 because of specific chemical properties of this radiopharmaceutical compound. The sensibility of this method is 80% and the specificity 97.8% .Intermediate filament keratin is regarded as a good marker for epithelial and mesothelial tumors. In the intracranial and intraspinal spaces keratin has been demonstrated only in the endocrine cells of the adenohypophysis, squamous epithelial islands in the pars tuberalis of the hypophysis, and in the choroid plexus epithelium. Since gliomas and meningiomas do not express keratin, immunohistochemistry essay for keratin provide an additional help for differentiating between metastatic melanomas and primary central nervous system tumors . The addition of monoclonal antibody immunocytology to conventional techniques significantly improves the sensitivity of CSF cytology. This is particularly useful in the diagnosis of “Neoplastic meningitis,” also called metastatic meningeal melanomatosis (MMM) a complication of malignant melanoma disease whose diagnosis rests on the demonstration of malignant cells within the CSF .The putative CSF tumor markers, fibronectin, and beta 2-microglobulin, are elevated significantly in MMM  but not in patients with solid cerebral metastases. A prominent increase in the IgM index, which reflects intrathecal B-cell stimulation, and rise of IgG index, interleukin-6, and tumor necrosis factor-alpha in MMM patients provide preliminary evidence for a local intrathecal immune response triggered by melanoma cell invasion of the subarachnoid space .Diagnosis of melanoma CNS metastases typically is made following the onset of clinical symptoms. Thus, more sensitive diagnostic approaches are needed to identify subclinical CNS metastases. Currently, standard cytological analysis of the cerebrospinal fluid (CSF) is limited by its poor sensitivity. A more sensitive assay was therefore developed using multiple reverse transcriptase-polymerase chain reaction (RT-PCR) markers.The CSF is collected and assessed by RT-PCR for three known melanoma-associated markers  to detect occult metastatic melanoma cells in the CSF. The correlation between CSF RT-PCR positivity of MART-1 and/or MAGE-3 and the development of CNS metastases is significant. Of the patients with positive CSF RT-PCR markers, 41% have either positive MRI and/or positive RT-PCR results .The optimal treatment of melanoma patients with CNS metastases depends on each situation. Often surgery, radiosurgery, whole brain radiotherapy and chemotherapy are used in combination to obtain longer remissions and optimal symptom relieve . PatientGamma Knife radiosurgery or surgical resection of CNS disease prior to chemotherapy improves survival versus delayed treatment in patients melanoma with melanoma brain metastases . RadiothSurgery of an isolated metastasis can lead to a long survival but brain lesions are frequently numerous and associated with an extracerebral diffusion . Complet Duration and quality of survival depend on the extent of metastatic disease and response to treatment. Treatment goals with WBI are palliation of symptoms and prolongation of life. Although brain metastases may be treated with surgery and/or stereotactic radiosurgery (SRS) when disease is limited to three lesions, treatment for patients with large or multiple metastases is limited to WBI. While formal response and survival analyses of the impact of WBI in melanoma metastases have not been reported, the estimated mean survival time for unselected patients with CNS metastases is only 2 to 4 months, with 1-year survival rates of less than 13%. This rates prevent the use of WBI as single therapeutic option. In a selected population of patients with limited CNS involvement, surgical resection alone or in combination with WBI appears to prolong mean survival. Overall survival is significantly improved in patients with multiple metastasis who receive adjunctive cranial irradiation versus those who had surgery alone . Therefore, adjunctive cranial irradiation is justified for melanoma patients who undergo surgical therapy for solitary brain metastases. Survival in patients presenting with solitary brain metastases is improved by a reduction of relapse in the brain as a component of failure by combined surgery and irradiation. In these patients survival will depend basically upon control of systemic disease Some authors even suggest the prophylactic whole brain irradiation for patients with melanoma that present high risk of metastases, once CNS metastases are sometimes the sole site of clinical relapse, and are frequently disabling . In accelerated irradiation regimens, the total tumor dose varies from 3000 to 4800 rad, and the overall treatment time from 1 to 2 weeks. This more aggressive form of treatment has demonstrated no significant improvement in the results from accelerated fractionation in the treatment of intracranial metastases. The result of the radiotherapy treatment did not depend on the site of the primary lesion, number of intracranial metastases, total dose, or the dose perfraction. There are, however, two subgroups not mutually exclusive, that benefit significantly from the accelerated fractionation: patients who had a complete resection of brain metastases, and those having no detectable extracranial metastases at the time of their treatment for intracranial metastases .Although in larger metastases the blood-brain barrier is leaky, lesions are resistant to many chemotherapeutic drugs. While systemic therapy for metastatic melanoma produces objective responses in 15% to 50% of patients, available drugs do not penetrate well into the CNS, and these patients rarely benefit from systemic therapy . Dacarbazine (DTIC) gives a mean response rate of 21% on visceral localizations but does not cross the blood brain barrier (BBB). Biological response modifiers like Interleukin 2 (Il2) leads to a 25% response rate in disseminated melanoma .The results indicate that systemic PCNU is unlikely to be more effective than other currently used chemotherapy in patients with malignant melanoma and CNS metastases . Moreoveomide include brain, lung, liver, lymph nodes and muscle. Patients tolerate treatment with termozolomide well and usually no dose reduction is necessary. However some few patients may present complications due to severe leucopoenia and thrombocytopenia (WHO grade 3 and 4). Thus, termozolomide represents a safe treatment option in patients with metastatic melanoma and poor prognosis [Temozolomide (TMZ) is a novel oral alkylating similar to dacarbazine (the most active single agent in primary melanoma) that have 100% oral bioavailability and considerable penetration of CNS tissue. TMZ has broad preclinical antitumoral activity that in melanoma is comparable to that of dacarbazine.Sites of remission of metastatic melanomas treated with temozolrognosis .omide reported is 7 months since beginning of therapy.Temozolomide (TMZ) may also be used in combination with interferon alpha-2b . The most common adverse events during the use of this association are fatigue, fever, nausea/emesis, anxiety, and diarrhea. Most toxicity is mild to moderate in severity. The primary dose-limiting toxicity is thrombocytopenia. The maximum tolerated dose is either TMZ 150 mg/m(2) plus IFN-alpha2b 7.5 MIU/m(2) or TMZ 200 mg/m(2) plus IFN-alpha2b 5.0 MIU/m(2). The pharmacokinetics of TMZ is not affected by coadministration of IFN-alpha2b , 34. TheFotemustine (muphoran), a new amino acid-linked nitrosourea, can give a response rate up to 28.2% in patients with cerebral metastases, and the increased survival of responding patients is significant , 29. All2 can be administered alone, with 1,3-bis(2-Chloroethyl)-1-nitrosourea (BCNU) or with bleomycin. Thirty percent of the patients have objective improvement in CT scans and about 13% of them have stabilization of disease. The median time to tumor progression for responding patients is 20 weeks. Neurological and retinal toxicity are potential complications of this therapy [Intracarotid cisplatin-based chemotherapy may be useful for palliation in selected patients with malignant melanoma and CNS metastases. Intracarotid cisplatin 40–75mg/m therapy .Interferon apparently is inactive against melanoma brain metastases, but does cause CNS symptoms .More recently, stereotactic radiosurgery (SRS) has been used as an effective local treatment for patients with CNS melanoma metastases. In several retrospective reports, treatment with SRS alone or in combination with whole brain irradiation (WBI) has demonstrated to prolong mean survival . As happGamma knife surgery (GKS) is effective in treating melanoma metastases in the brain. It appears that the radiobiology of a single high dose overcomes the radioresistance barrier, yielding better results than fractionated radiation. Twenty-four percent of the lesions treated with GKS disappear, 35% shrink, 23% remain unchanged, and 18% increasing size. No undue radiation-induced changes are observed in the surrounding brain. No deaths or neurological morbidity related to GKS is observed. The mean survival time calculated is 10.4 months from the time of treatment with GKS. Solitary brain lesions and lack of visceral metastases are statistically predictive of a better prognosis .Boron neutron capture therapy (BNCT) is yet an incipient therapy method that has shown good results in rats experiments, mainly if combined by either Cereport (RMP-7) mediated modulation of blood-brain barrier (BBB) permeability or hyperosmotic mannitol-induced BBB disruption using borono-phenylalanine (BPA) as the capture agent. Optimizing the delivery of BPA by means of intracarotid injection combined with opening the BBB by infusing Cereport or a hyperosmotic solution of mannitol significantly enhance survival times and produce long-term cures in animals with CNS melanoma metastasis. These observations are relevant to future clinical studies using BNCT for the treatment of intracerebral melanoma .Prognosis for patients with melanoma brain metastasis is still poor with a mean survival time of 6 months after diagnosis , 5, 42. Most of the symptoms of CNS melanoma metastasis are unspecific and depend on localization of the lesion. CT scan is recommended as the primary exam in case of suspected cases. MRI must be used to study more precisely the characteristics of the lesions in order to embase the surgical plane. It is important that the clinician suspect of the possibility of CNS metastasis in all patients with new neurological signs and previous primary melanoma lesion. These patients deserve a careful clinical analysis and further exams for investigation .Optimal treatment of melanoma patients with CNS metastases depends on the objective situation. Often surgery, radiosurgery, whole brain radiotherapy and chemotherapy are used in combination to obtain longer remissions and optimal symptom relieve . Patient


Posterior circulation strokes represent approximately 20% of all ischemic strokes , 2. In c A posterior circulation (PC) stroke is classically defined by infarction occurring within the vascular territory supplied by the vertebrobasilar (VB) arterial system. The vertebral arteries (VAs) arise from the right and left subclavian arteries and travel cranially through the transverse foramina of the cervical vertebrae. When reaching the foramen magnum, they pierce the dura mater to start their intracranial course. Both VAs join at the pontomedullary junction forming the basilar artery (BA). The VAs are divided into four segments. V1 is the most proximal segment from the VA origin to point of entry into the initial foramen transversarium, usually at the sixth or seventh cervical vertebral body. The segment occupying the course of the VA from the initial to last transverse foramen is known as V2. The V3 segment begins after the artery exits the foramen transversarium of the axis (C2) and arches behind the atlas (C1) before entering the cranium. The most distal segment representing the intracranial portion of the VA after entering the dura mater is the V4 segment. The BA travels rostral along the ventral medulla and basis pontis until it bifurcates into the right and left posterior cerebral arteries (PCAs) at the pontomesencephalic junction. Segments of the vertebral artery and branches of the vertebrobasilar system are depicted in Figure Anatomical variations of the PC are frequent but commonly asymptomatic. Familiarity with these variants is important as not to mistake them as pathological findings. Although typically discovered incidentally, they are infrequently related to intracranial vascular pathology.Asymmetric VAs occur in over two-thirds of individuals , 5 FiguA and an The artery of Percheron is where a single thalamic perforating artery arises from the proximal PCA (P1 segment) between the BA and PCOM and supplies the rostral mesencephalon and both paramedian thalami , and PCA branches . AC stroThe most common intracranial site of atherostenosis is the BA, followed by the ICAs, MCAs, VAs, PCAs, and ACAs. Intracranial stenosis is more prevalent among African-American and Asian individuals and may account for a higher rate of strokes in these populations compared to Caucasians Figure B. Neurovin situ thrombosis, or artery-to-artery thromboembolism. Mild intracranial disease may have minimal effect on cerebral hemodynamics. As the stenosis increases, reflex vasodilation due to inadequate or failing collateral circulation occurs to increase cerebral blood volume (CBV) and preserve normal cerebral blood flow (CBF), and oxygen extraction fraction will increase as CBF further deteriorates. Failure of these compensatory mechanisms is known as “misery perfusion”  was the most common etiology (40%) and a cardiac source of embolism was reported in 24%. Distal PC territory infarctions were most common followed by the middle territory . In contSimilarities and differences in both registries exist. The prevalence of hypertension, diabetes, hyperlipidemia, and smoking were similar in both registries. Over 80% of both cohorts were investigated with magnetic resonance imaging (MRI) and magnetic resonance angiography (MRA). Compared to HSR, the NEMC-PCR included a higher proportion of men 63 vs. 56%) and patients were slightly younger (mean age 61 vs. 63 years). NEMC-PCR patients were predominantly Caucasian (84%) and only a minority was Asian (9.5%)  was only performed in 37.5%, transesophageal echocardiography (TEE) in 14.1% in contrast to the NEMC-PCR, and VA origins were not routinely evaluated . The lowVertebrobasilar dolichoectasia (VBD) refers to dilatation, elongation, and tortuosity of the BA. The anatomy is highly variable. Asymptomatic patients may be found to have VBD incidentally on neuroimaging while other patients present with vertebrobasilar territory ischemia . Risk faCervical artery dissections (CADs) may occur spontaneously or result from major or minor cervical trauma . In younOther less common causes of ischemia with predilection for the PC circulation include subclavian steal syndrome, giant-cell arteritis, and Fabry disease. Mitochondrial encephalopathy, lactic acidosis, and stroke like episodes (MELAS), migraines, and posterior reversible encephalopathy syndrome (PRES) also have a predilection for the PC . ReversiIn clinical practice, not all PC stroke presentations are classic. Many patients present with signs and symptoms of multifocal PC infarctions. Moreover, the PC is rich in potential collateral support and clinical manifestations of BA ischemia may be highly variable. Symptoms associated with PC strokes such as diplopia, visual field defects, dysphagia, vertigo, alteration in consciousness, or hearing loss may aid with localization. PC strokes have fewer cortical findings and relatively small lesions can cause significant deficits as compared to AC stroke due to the close proximity of major afferent and efferent tracts and cranial nerve nuclei in the brainstem. Table Common presenting symptoms of PC stroke include vertigo, imbalance, unilateral limb weakness, slurred speech, double vision, headache, nausea, and vomiting. Exam findings include unilateral limb weakness, gait ataxia, limb ataxia, dysarthria, and nystagmus. Infarcts involving the proximal PC territory may cause dysphagia due to pharyngeal weakness, nausea, vomiting, and Horner’s syndrome. Infarcts involving the middle territory are often associated with limb weakness and nuclear facial palsy. Distal territory infarctions are commonly associated with decreased appendicular sensory loss, lethargy, and visual field defects . PatientTypical of stroke, symptoms are sudden and maximal at onset. Vertebrobasilar insufficiency (VBI) refers to PC symptomatology due to hemodynamic failure resulting in ischemia. A steno-occlusive lesion at any level due of the VA due to any cause may lead to VBI and a physical examination alone is insufficient for diagnosis . A detaiVertebrobasilar TIA may precede PC stroke in approximately one-fourth of cases . EpisodeHistory of unilateral arm pain with exertion associated with VBI symptomatology may suggest proximal subclavian artery stenosis causing reversal of normal flow within the ipsilateral vertebral and basilar arteries, known as the subclavian steal syndrome . A bloodVertebrobasilar symptoms or syncope with head-turning known as rotational vertebrobasilar insufficiency or bow hunter’s syndrome is rare. This is most commonly caused by compression of the vertebral artery due to cervical bony osteophyte in older patients, C1–C2 hypermobility, or even neck muscle hypertrophy in younger patients , 49.Dizziness, albeit descriptive, is a non-specific term that is commonly used interchangeably with vertigo by patients. Moreover, patients commonly use dizziness to describe feeling faint or light-headed. Nearly 7.5 million patient visits per year in the ambulatory setting are due to dizziness as the presenting symptom , 51. In In contrast, in 240 patients clinically diagnosed with vestibular neuritis due to vertigo lasting >24 h without accompanying neurological symptoms and a normal neurologic examination, cerebellar infarctions were found in 10% of patients. Almost all infarctions were within the PICA territory . HoweverQualitative descriptions may be less dependable than estimates of the duration and situations that preceded or provoked the event . Brief, Ischemia associated with vertigo is often accompanied by other brainstem or cerebellar signs and symptoms . ProvocaIn patients presenting with acute vestibular symptoms , the HINTS method, a three-step bedside evaluation composed of horizontal head impulse testing, nystagmus evaluation, and testing of skew can be easily performed in about 1 min. A negative  head impulse, alternating nystagmus, and presence of skew (refixation on cover–uncover testing) has a sensitivity and specificity of 100 and 96%, respectively, for detecting stroke. In addition, the presence of skew deviation is highly specific for brain stem dysfunction in patients with the aforementioned symptoms .Gait ataxia may be central, vestibular, or sensory in origin. It is almost always present in PC stroke involving the brainstem and cerebellum  and nearHeadaches occur commonly in the general population due to myriad etiologies and are non-specific symptoms in many cases. The frequency of headache with stroke is approximately 27%, and increases to 46–70% when the PC is involved . HeadachBrain computed tomography (CT) is typically performed as the initial imaging modality for patients presenting with acute stroke symptoms. Unfortunately, CT provides suboptimal visualization of the posterior fossa structures due to obscuration by artifacts produced by the bony structures of the cranial base and early ischemic changes may not be visible. In contrast, MRI provides better visualization of the soft tissue structures  and is sSubtle hypodensities, loss of gray–white matter differentiation and sulcal effacement have been used to asses for signs of early ischemia on non-contrast head CT. The Alberta Stroke Program Early CT Score (ASPECTS) provides a reliable and standardized topographic assessment of acute ischemic changes in the MCA territory with predictive value , 63. In Despite the superior sensitivity of MRI-DWI in detection of PC stroke compared to CT, MRI can rarely be negative for small infarctions . In one Presence of a hyperdense BA sign in the setting of acute PC stroke may be indicative of thrombosis. This finding has been shown to be 71% sensitive, 98% specific, and 94% accurate for BA occlusion with a positive predictive value of 83%, negative predictive value of 95%, and a strong predictor of short- and long-term outcome . The golIdeal treatment of symptomatic vertebrobasilar stenosis and indications for invasive treatments remains a topic of debate as randomized controlled trials data are lacking. Patients with symptomatic vertebrobasilar stenosis, defined as VA or BA stenosis >50% by CTA or contrast-enhanced MRA have a threefold higher 90-day risk of stroke or TIA after a first event as compared to patients without. Stroke risk is highest within the first 2 weeks and patients with intracranial stenosis harbor an even higher risk of early recurrence, up to 33% as compared to 16.2% for extracranial stenosis  have notIn the Carotid and Vertebral Artery Transluminal Angioplasty Study (CAVATAS), only eight patients underwent endovascular angioplasty with or without stenting precluding meaningful conclusion about the utility of endovascular intervention within the VA . A recenIn the stenting of symptomatic atherosclerotic lesions in the vertebral or intracranial arteries (SSYLVIA) trial, the 30-day perioperative stroke rate for intracranial stenting was 6.6% . RestenoSpecific guidelines for treatment of VBD are lacking. Management is typically conservative with use of antiplatelet agents or anticoagulants. However, antithrombotic use may be associated with an increased risk of intracerebral hemorrhage . VentricPatients with PC stroke and atrial fibrillation should be managed with long-term anticoagulation. Available oral agents include vitamin K antagonist therapy with warfarin , direct thrombin inhibitors such as dabigatran, and factor Xa inhibitors such as apixaban, edoxaban, and rivaroxaban . PatientAcute treatment options for PC stroke include IV recombinant tissue-plasminogen activator (IV-rt-PA), intra-arterial fibrinolysis, and endovascular thrombectomy. The acute management of ischemic stroke is primarily governed by time from last known well and comorbid conditions. Within the therapeutic treatment window, stroke severity based on the NIHSS play an important role in decision-making. However, this measure has limitations in PC stroke. Higher points in AC stroke are seen with cortical findings and motor deficits, in contrast to fewer points assigned to cranial nerve deficits and ataxia which may occur without weakness in PC strokes , 90. PatOnly 5% of the National Institute of Neurological Disorders and Stroke (NINDS) Study that led to U.S. Food and Drug Administration approval of alteplase for acute ischemic stroke had PC stroke  and dataBasilar artery occlusion (BAO) is a neurological emergency that carries >80% fatality rate without treatment , 97 promThe use of intra-arterial (IA) thrombolytics in PC stroke has been previously studied , 101. FaRecanalization has a strong association with clinical outcome. A large meta-analysis of 53 studies with reported recanalization rates including 2066 patients demonstrated a four- to fivefold increase in the odds of a good functional outcome and similar decrease in the odds of death in patients with successful recanalization . In addiRecent randomized controlled trials have not clarified the role of endovascular therapy for acute ischemic stroke in general and PC stroke in particular. The interventional management of stroke III (IMS III) trial, showed no overall benefit of endovascular rescue therapy after IV-rt-PA compared to IV-rt-PA alone in 656 randomized patients with ischemic stroke and NIHSS ≥10. However, older generation thrombectomy devices were employed, onset-to-treatment times were long, and only 2% of patients had PC stroke. A favorable trend was seen for patients receiving IV-rt-PA within 2 h of symptom and endovascular rescue therapy within 90 min of IV-rt-PA initiation . The SYNThe main mechanism of stroke after cervical arterial dissection is thromboembolism rather than hemodynamic compromise. This prompts consideration regarding antithrombotic selection for recurrent stroke prevention . AntithrCerebellar infarctions can rapidly swell due to cytotoxic edema within the tight constraints of the non-yielding posterior fossa and lead to obstructive hydrocephalus or fatal herniation if not adequately managed in a timely fashion –119 FigD–F. PatiSeveral anatomical and clinical differences distinguish PC infarctions from those in the AC and have practical implications. Vascular anatomical variation may affect stroke severity and presenting signs and symptoms. Large vessel atherostenosis is common but cardioembolic strokes, dissections, and other etiologies are known to affect the PC. Quantitative measures such as the NIHSS alone may not suffice and clinical signs and symptoms such as dizziness, vertigo, and ataxia may be diagnostic pitfalls. MRI is superior to CT for imaging an acute PC infarct but may rarely miss a small infarct early on. A variety of non-invasive vascular imaging modalities are available to aid the etiologic work-up and have replaced conventional catheter angiography in many circumstances. IV-rt-PA is the recommended first-line treatment for eligible patients. The role of endovascular acute revascularization therapy remains uncertain at present. The cornerstone of recurrent stroke prevention includes risk factor control primarily blood-pressure lowering, lipid-lowering with statins, and antithrombotic therapy. Aggressive medical management has been shown to be superior to endovascular angioplasty and stenting in patients with a range of intracranial atherostenosis including lesions in the PC.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The extent to which mass extinctions influence body size evolution in major tetrapod clades is inadequately understood. For example, the ‘Lilliput effect,’ a common feature of mass extinctions, describes a temporary decrease in body sizes of survivor taxa in post-extinction faunas. However, its signature on existing patterns of body size evolution in tetrapods and the persistence of its impacts during post-extinction recoveries are virtually unknown, and rarely compared in both geologic and phylogenetic contexts. Here, I evaluate temporal and phylogenetic distributions of body size in Permo-Triassic therocephalian and cynodont therapsids (eutheriodonts) using a museum collections-based approach and time series model fitting on a regional stratigraphic sequence from the Karoo Basin, South Africa. I further employed rank order correlation tests on global age and clade rank data from an expanded phylogenetic dataset, and performed evolutionary model testing using Brownian (passive diffusion) models. Results support significant size reductions in the immediate aftermath of the end-Permian mass extinction (ca. 252.3 Ma) consistent with some definitions of Lilliput effects. However, this temporal succession reflects a pattern that was underscored largely by Brownian processes and constructive selectivity. Results also support two recent contentions about body size evolution and mass extinctions: 1) active, directional evolution in size traits is rare over macroevolutionary time scales and 2) geologically brief size reductions may be accomplished by the ecological removal of large-bodied species without rapid originations of new small-bodied clades or shifts from long-term evolutionary patterns. Body size is an important biological trait that affects organismal fitness and imparts a strong influence on ecological, physiological, and life history attributes of species Lystrosaurus Assemblage Zone, Karoo Basin of South Africa Though originally reserved for within-lineage size decreases in survivor taxa Lystrosaurus Assemblage Zone (hereafter ‘AZ’) using non-phylogenetic time series and statistical comparisons of size distributions between temporally successive eutheriodont assemblages. Second, I chose to implement time series modeling and rank correlation tests to evaluate whether reductions, if present, signified transient shifts or longer-term directional size shifts both across the extinction and during pre-extinction intervals. A transient shift following mass extinction (an expectation of Lilliput-type patterns) would be indicated by short-term shifts in stratigraphic series without a significant mean step change across the entire sampled interval .Non-random extinction patterns may be discernible with regard to some organismal traits, especially when studied in the context of existing patterns of trait evolution during background intervals. Non-phylogenetic investigations of selectivity in Cretaceous-Paleogene marine invertebrates have suggested that mass extinction may bring about changes in selectivity patterns that reset the stage for biotic recoveries. This departure from normal (background) selectivity patterns has been termed “non-constructive selectivity” Lystrosaurus AZ. Recent assessments of geologic controls on paleodiversities have found potential associations between outcrop area and taxonomic richness of pre-extinction assemblages, but the outlying Lystrosaurus AZ exhibits unexpectedly modest richness that is unexplainable by outcrop area alone Lystrosaurus AZ. While taphonomic and collecting biases are important considerations, I suggest that they would have had limited impacts on diversity and body size shifts observed in the Lystrosaurus AZ.Sampling of non-isotaphonomic intervals and unevenness of sampling may bias the quality and size controls of the present sample. These can include but are not restricted to available outcrop area, collection intensity, and sediment coarseness and sorting, which should be considered given the comparatively small sizes observed in the Triassic Size shifts in post-extinction faunas, if present, may have been underpinned to some extent by stochastic processes of trait evolution. Long-term negative shifts observed throughout the sampled interval may indicate background patterns of trait evolution . Size shifts may alternatively represent rapid diversifications of new clades whose success was associated with the trait in question . Rapid diversification of small-bodied taxa following extinction is one explanation for Lilliput-type patterns, although few examples have been demonstrated Eodicynodon, Tapinocephalus, Pristerognathus, Tropidostoma, Cistecephalus, Dicynodon, Lystrosaurus, and Cynognathus AZs. Additional global data, collected to distinguish non-regional geologic and phylogenetic trends in all sampled Permian and Triassic taxa, covered a wider stratigraphic interval so that late Middle Triassic (Ladinian) and Late Triassic  global standard stages were also included as distinct bins. The sample was limited to Permian and Triassic genera only, and to specimens having well-preserved skulls for complete skull length measurements. Several Early Jurassic nonmammaliaform cynodonts from the Karoo  and those lacking complete skulls were therefore excluded from the dataset. The resulting database included 347 complete cranial specimens and 72 species of Permian through Triassic therocephalians and cynodonts from the Karoo Basin and contemporaneous basins. Sources for occurrence data, including voucher specimens and absolute dates, are provided in I assembled a database of specimen measurements and stratigraphic occurrences for 72 species of Permian and Triassic therocephalians and cynodonts. The majority of measurements were taken directly from museum specimens, but data for some non-African taxa that were inaccessible during the study period were taken from the literature. Permission to access collections was obtained from all visited institutions prior to the study period. By incorporating data from museum specimens, I was able to assess variation within species, including maximum sizes, rather than representative samples from the literature. A rigorous, collections-based survey therefore allows previous literature-based assessments of body size evolution in Permo-Triassic therapsids to be improved upon From the complete database, 57 measurements were retained for time series analysis of the Karoo assemblages, and 72 measurements were retained for rank order analyses of the global dataset representing the maximum basal skull lengths of the 72 sampled taxa from the stratigraphic interval of their first appearance . Basal skull length (BSL), measured from the tip of the snout to the occiput in millimeters (mm), was chosen as a proxy for body size for several reasons. Firstly, many of the fossil taxa examined here are best represented by cranial material, which is more accessible than postcranial elements and is relatively diagnostic for nonmammalian therapsids (contributing to the accuracy of specimen identifications). Moreover, investigation of BSL provides direct comparisons to other published studies using similar proxies. Skull length also correlates well with other linear skeletal measurements in therapsids such as femur length Cistecephalus and Dicynodon AZs) and earliest Triassic (Lystrosaurus AZ) were drawn from equivalent distributions using a Kolmogorov-Smirnov (K-S) test. Then, I evaluated the likelihood that observed negative shifts in size distributions (if present) represent a random evolutionary walk  by assessing the record of body size evolution over eight stratigraphic bins of the Karoo succession. Mean lnBSL was estimated for each of eight stratigraphic bins by taking the average of the maximum known lnBSL for each taxon within a given bin, therefore representing a per taxon average . In contrast to the rank order correlation tests, which examined only size at first appearance (discussed below), taxa whose ranges spanned multiple assemblage zones were included in each bin in which they occurred in order to capture more fully the variance within each bin. Mean sizes were estimated in each bin for all eutheriodonts, as well as separately for therocephalians and cynodonts in order to evaluate the pervasiveness of potential trends among the therapsid subclades.Time series data represent a sequence of observations repeated over elapsed time, and can be conceptualized in a paleontological context as ancestor-descendant relationships  Lystrosaurus AZ, then longer-term size reductions should not be manifested in the Karoo time series, with non-directional models best fitting the data . If, on the other hand, body size evolution was driven by longer-term trends beginning in the Permian, then a directional model with a negative mean step change should best fit the data, thereby refuting the rapid and short-term perturbation predicted by the Lilliput effect.Mean lnBSL and disparity (as measured by variance) were estimated from the database for each Karoo biozone (average bin duration ≈3 Myr) and imported into the ‘R’ version 2.15.1 statistical environment using the ‘paleoTS’ package The relative fit of the three models  was assessed using Akaike's information criterion corrected for small sample sizes (AICc). AICc is an information loss metric that is used to compare models both through their goodness-of-fit and model complexity . AICc diminishes the problem of overfitting complex models, and is widely used for small paleontological and neontological datasets. Akaike weights (‘AW’) are estimated by converting the individual AICc scores for each candidate model into a proportional value representing the model's relative support ; the model with the highest weight represents that with the least information loss . A more complete description of AICc and its application to time series and phylogenetic model fitting can be found in Hunt and Carrano ρ) and Kendall's tau (τ) rank order tests. These tests allowed potentially positive or negative trends in the global dataset to be evaluated as a monotonic function . Importantly, unlike their parametric equivalent, Pearson's product moment correlation, the tests are robust even when the association is not linear. Three separate data subsets were analyzed to evaluate directional size trends in eutheriodonts (N = 72) and therocephalian (N = 37) and cynodont (N = 35) subclades from the Middle Permian through Late Triassic. For Eutheriodontia and its subclades, I evaluated ρ and τ coefficients for negative trends in two ways, assessing 1) data subsets that included taxa only having first appearances in the Permian and 2) the total Permian-Triassic dataset. By examining Permian-only and excluding Triassic originations, I was able to assess the extent to which general trends in the total Permian-Triassic dataset were influenced by new size distributions of post-extinction taxa (versus pre-existing or ‘background’ patterns of trait evolution). Trends were considered only weakly significant if both coefficients were negative but non-significant for one of the tests.Rank order statistics provide additional tests for identifying directional trends in ranked paleontological data  Lystrosaurus AZ. Visual inspection of the Karoo time series data suggests a transient decrease in mean size from the Late Permian Dicynodon AZ to the Lystrosaurus AZ, which rebounded subsequently in the Cynognathus AZ . A binomial test on the Karoo dataset using the binom.test function of paleoTS Lystrosaurus AZ therefore cannot be attributed to a long-term directional trend when evaluated as a time series.My evaluation of stratigraphic  patterns failed to identify long-term negative trends in eutheriodont body size evolution, instead showing a short-term reduction in mean sizes in the Triassic athus AZ . KolmogoRank order correlations between age rank and size at first appearance, on the other hand, may indicate a weak negative trend for some groups such as therocephalians, which originated at their largest sizes in the Permian , Table 2Cladistic data were also ranked to permit the evaluation of directional trends in size data with respect to relative branching events in a reference cladogram . I constAncestor-descendant tests were performed as an extension of the CR analyses in order to assess tendencies toward negative versus positive size changes in the phylogenetic tree between reconstructed ancestor and descendant nodes ymmeTREE version 1.1 was used to evaluate the topological distribution of taxic diversity across branches in the reference phylogeny  and to identify regions of the tree that contribute to significant imbalance in diversity. This operation was performed by calculating the Δ1 and Δ2 statistics, functions of two likelihood ratios that represent the probability of observing a shift in diversification rate along the internal branch of a three-taxon tree . For each node, the ratios compared the likelihood of yielding the observed distribution of diversity under an equal-rates Markov branching model to that under a two-rate model. Importantly, the inclusion of non-contemporaneous clades sampled from disjunct stratigraphic bins may violate an assumption of the equal-rates Markov model by heterogeneously sampling diversity through time Tropidostoma, Cistecephalus, Dicynodon, Lystrosaurus, and Cynognathus AZs. Following Botha-Brink and Angielczyk ymmeTREE to identify diversification rate shifts in post-extinction clades  or in clades that diversified before the extinction .To assess the hypothesis that shifts in mean body size represented post-extinction radiations of small-bodied eutheriodont clades, a phylogeny was constructed using all studied taxa in date.phylo function (http://www.graemetlloyd.com/methdpf.html) T), Ornstein-Uhlenbeck (OU), and early burst Brownian motion (EB). BM models evolution as a process of passive diffusion ; BMT, models evolution as a generalized random walk having a positive or negative mean step change ; OU models trait evolution as a constrained process approaching some optimal value (sometimes equated to ‘stabilizing selection’); and EB approximates a Brownian process in which the rate of trait evolution exponentially decreases (‘decays’) over time. Models were fitted to the data using ‘geiger,’ and likelihood ratio tests were performed to determine whether multiparameter models should be selected over the null BM model. Goodness-of-fit tests implementing AICc were also used to evaluate the candidate models as in the time series analysis.Finally, stochastic models of Brownian evolution were fitted to observed patterns of body size evolution in a comprehensive global phylogeny of eutheriodonts, permitting a marriage of temporal and topology-based methods. Phylogenetic model fitting was performed in R using ‘ape’ and ‘geiger’ packages ρ = −0.622, p<0.001; τ = −0.484, p<0.001) and a weakly negative correlation in cynodonts . The same result is true for therocephalians whether analyzed with or without Triassic taxa, indicating that size reductions began in relatively shallowly-nested nodes in the tree  and continued into the Triassic. Correlations with patristic distance have lower statistical support, but are still significant for Permian-only and Permian-Triassic therocephalians indicating the influence of therocephalians on the overall pattern. I carried out ancestor-descendant tests following the method of Carrano Cladistically-informed analyses were better able to detect long-term body size trends than stratigraphic data alone ; Table 3Lystrosaurus or Cynognathus AZ diversity partitions, suggesting that either sampling is too poor to detect a shift or that the impact of the extinction on diversification rates in eutheriodonts was limited. The only significant diversification rate shift occurred at the early-Late Permian divergence of Eutherocephalia . Accordingly, no significant rate shifts were detected in tree topologies constructed from Triassic cephalia , a resulT) and ‘stabilizing’ (OU) models fit poorly when compared to other candidate models of passive, Brownian motion evolution (BM and EB). Likelihood ratio tests and comparisons of Akaike weights indicate that EB best approximates the mode of body size evolution across the global dataset of eutheriodonts . An unexpected result is the protracted and increasingly constrained evolutionary rates in late Middle to Late Triassic cynodonts. The factors influencing their constrained phenotypic evolution are beyond the scope of the present analysis, but may include intrinsic factors  or extrinsic selective pressures (competition with larger-bodied archosaurs or other emerging clades) that merit future study.Body size evolution before and immediately after the end-Permian extinction cannot demonstrably be classified as active or “driven” sensu McShea Lystrosaurus AZ taxa and clades have Permian ghost lineages  within-taxon size reductions in relict lineages, 2) extinction of large taxa, and/or 3) post-extinction radiations of small taxa lineages , 8, implMoschorhinus and Lystrosaurus), but more general shifts toward smaller terrestrial vertebrate taxa were likely accentuated by the ecological removal of large-bodied Permian species . Recent surveys of bone histology in these Permian-Triassic survivor taxa provide little evidence that Triassic therapsids grew more slowly than their Permian predecessors, and in some cases they may have grown more quickly or over shorter durations This interplay of Lilliput mechanisms parallels that of some Permian-Triassic marine invertebrate clades in some respects, but differs in others. Global surveys of marine gastropods suggest strong within-lineage size decreases as well as size-selective extinction Appendix S1List of institutional abbreviations.(PDF)Click here for additional data file.Appendix S2Outcrop area and sampled diversity of Karoo Basin therocephalians and cynodonts.(PDF)Click here for additional data file.Appendix S3Age and clade rank data for therocephalian and cynodont subclades. Basal skull length measurements (BSL) are in mm.(PDF)Click here for additional data file.Dataset S1Phylogenetic methods and supporting NEXUS file.(PDF)Click here for additional data file.


Approximately 30% of AIDS-related lymphomas (ARL) are associated with infection by the EBV, and about 4% by the KSHV/HHV-8. It is likely that if other lymphomagenic pathogens exist, these associations would occur in the context of ARL. The advent of high throughput sequencing provides a unique opportunity to address this question. High throughput sequencing, followed by computational subtraction of human sequences was used for enrichment of candidate pathogenic sequences.PathSeq, which carries out several subtraction steps involving alignments to i) human genome sequence databases; ii) human transcriptome sequence databases; and iii) other vertebrate sequence databases. Residual sequence reads were then compared with microbial databases, either individually or as part of de novo assembled contigs.Eleven primary tissues of ARL have been used to generate cDNA libraries. Out of these six were frozen specimens and five were formalin-fixed paraffin-embedded (FFPE). These libraries were subjected to high throughput Illumina sequencing to generate 30-60 million 76bp paired-end sequence reads per sample. Quality filtered reads were analyzed using our automated pipeline, Using both frozen and formalin-fixed paraffin-embedded (FFPE) tissues, we have identified unique sequences previously unassociated with hematological cancers and inflammatory diseases. In addition, our pathogen discovery pipeline works with both transcriptome and whole genome sequencing (WGS) data, and it is applicable to data across all high throughput sequencing platforms. Most notably, we are able to detect as low as 1 viral sequence per billion total sequences for WGS data, a sign of the sensitivity of our method. Among the known pathogens, we found 12423 sequences corresponding to EBV in the one case where the presence of this virus was also documented by EBER in situ hybridization. Three additional cases that were EBER-negative revealed EBV sequences, in the range of 3 to 351 reads, suggesting that the virus was present in tumor-infiltrating cells, rather than in the lymphoma. Eight cases had HIV sequences ranging from 1 to 403 reads, and one case had a single read corresponding to KSHV.PathSeq, for pathogen discovery in both frozen and FFPE tissues using a high throughput sequencing-based computational subtraction process. The presence of >10,000 reads in the known EBV-positive case confirms the effectiveness of the method. Specific EBV and HIV sequences were seen emanating from tumor-infiltrating cells that will shed light on expression patterns of these viruses in this cellular compartment.We have developed an integrated pipeline,