Novel, objective, multivariate biomarkers composed of plasma amino acid profiles for the diagnosis and assessment of inflammatory bowel disease

Background

Inflammatory bowel disease (IBD) is a chronic intestinal disorder that is associated with a limited number of clinical biomarkers. In order to facilitate the diagnosis of IBD and assess its disease activity, we investigated the potential of novel multivariate indexes using statistical modeling of plasma amino acid concentrations (aminogram).

Methodology and Principal Findings

We measured fasting plasma aminograms in 387 IBD patients (Crohn''s disease (CD), n = 165; ulcerative colitis (UC), n = 222) and 210 healthy controls. Based on Fisher linear classifiers, multivariate indexes were developed from the aminogram in discovery samples (CD, n = 102; UC, n = 102; age and sex-matched healthy controls, n = 102) and internally validated. The indexes were used to discriminate between CD or UC patients and healthy controls, as well as between patients with active disease and those in remission. We assessed index performances using the area under the curve of the receiver operating characteristic (ROC AUC). We observed significant alterations to the plasma aminogram, including histidine and tryptophan. The multivariate indexes established from plasma aminograms were able to distinguish CD or UC patients from healthy controls with ROC AUCs of 0.940 (95% confidence interval (CI): 0.898–0.983) and 0.894 (95%CI: 0.853–0.935), respectively in validation samples (CD, n = 63; UC, n = 120; healthy controls, n = 108). In addition, other indexes appeared to be a measure of disease activity. These indexes distinguished active CD or UC patients from each remission patients with ROC AUCs of 0.894 (95%CI: 0.853–0.935) and 0.849 (95%CI: 0.770–0.928), and correlated with clinical disease activity indexes for CD (r_s = 0.592, 95%CI: 0.385–0.742, p<0.001) or UC (r_s = 0.598, 95%CI: 0.452–0.713, p<0.001), respectively.

Conclusions and Significance

In this study, we demonstrated that established multivariate indexes composed of plasma amino acid profiles can serve as novel, non-invasive, objective biomarkers for the diagnosis and monitoring of IBD, providing us with new insights into the pathophysiology of the disease.     

Interferon-gamma release assay (modified QuantiFERON) as a potential marker of infection for Leishmania donovani, a proof of concept study

Background

In areas endemic for visceral leishmaniasis (VL), a large number of infected individuals mount a protective cellular immune response and remain asymptomatic carriers. We propose an interferon-gamma release assay (IFN-γRA) as a novel marker for latent L. donovani infection.

Methods and Findings

We modified a commercial kit (QuantiFERON) evaluating five different leishmania-specific antigens; H2B, H2B-PSA2, H2B-Lepp12, crude soluble antigen (CSA) and soluble leishmania antigen (SLA) from L. donovani with the aim to detect the cell-mediated immune response in VL. We evaluated the assay on venous blood samples of active VL patients (n = 13), cured VL patients (n = 15), non-endemic healthy controls (n = 11) and healthy endemic controls (n = 19). The assay based on SLA had a sensitivity of 80% (95% CI = 54.81–92.95) and specificity of 100% (95% CI = 74.12–100).

Conclusion

Our findings suggest that a whole-blood SLA-based QuantiFERON assay can be used to measure the cell-mediated immune response in L. donovani infection. The positive IFN-γ response to stimulation with leishmania antigen in patients with active VL was contradictory to the conventional finding of a non-proliferative antigen-specific response of peripheral blood mononuclear cells and needs further research.     

Post-streptococcal auto-antibodies inhibit protein disulfide isomerase and are associated with insulin resistance

Post-streptococcal autoimmunity affects millions worldwide, targeting multiple organs including the heart, brain, and kidneys. To explore the post-streptococcal autoimmunity spectrum, we used western blot analyses, to screen 310 sera from healthy subjects with (33%) and without (67%) markers of recent streptococcal infections [anti-Streptolysin O (ASLO) or anti-DNAse B (ADB)]. A 58 KDa protein, reacting strongly with post-streptococcal sera, was identified as Protein Disulfide Isomerase (PDI), an abundant protein with pleiotropic metabolic, immunologic, and thrombotic effects. Anti-PDI autoantibodies, purified from human sera, targeted similar epitopes in Streptolysin O (SLO, P51-61) and PDI (P328-338). The correlation between post-streptococcal status and anti-human PDI auto-immunity was further confirmed in a total of 2987 samples (13.6% in 530 ASLO positive versus 5.6% in 2457 ASLO negative samples, p<0.0001). Finally, anti-PDI auto-antibodies inhibited PDI-mediated insulin degradation in vitro (n = 90, p<0.001), and correlated with higher serum insulin (14.1 iu/ml vs. 12.2 iu/ml, n = 1215, p = 0.039) and insulin resistance (Homeostatic Model Assessment (HOMA) 4.1 vs. 3.1, n = 1215, p = 0.004), in a population-based cohort. These results identify PDI as a major target of post-streptococcal autoimmunity, and establish a new link between infection, autoimmunity, and metabolic disturbances.     

Analysis of clonal type-specific antibody reactions in Toxoplasma gondii seropositive humans from Germany by peptide-microarray

Background

Different clonal types of Toxoplasma gondii are thought to be associated with distinct clinical manifestations of infections. Serotyping is a novel technique which may allow to determine the clonal type of T. gondii humans are infected with and to extend typing studies to larger populations which include infected but non-diseased individuals.

Methodology

A peptide-microarray test for T. gondii serotyping was established with 54 previously published synthetic peptides, which mimic clonal type-specific epitopes. The test was applied to human sera (n = 174) collected from individuals with an acute T. gondii infection (n = 21), a latent T. gondii infection (n = 53) and from T. gondii-seropositive forest workers (n = 100).

Findings

The majority (n = 124; 71%) of all T. gondii seropositive human sera showed reactions against synthetic peptides with sequences specific for clonal type II (type II peptides). Type I and type III peptides were recognized by 42% (n = 73) or 16% (n = 28) of the human sera, respectively, while type II–III, type I–III or type I–II peptides were recognized by 49% (n = 85), 36% (n = 62) or 14% (n = 25) of the sera, respectively. Highest reaction intensities were observed with synthetic peptides mimicking type II-specific epitopes. A proportion of the sera (n = 22; 13%) showed no reaction with type-specific peptides. Individuals with acute toxoplasmosis reacted with a statistically significantly higher number of peptides as compared to individuals with latent T. gondii infection or seropositive forest workers.

Conclusions

Type II-specific reactions were overrepresented and higher in intensity in the study population, which was in accord with genotyping studies on T. gondii oocysts previously conducted in the same area. There were also individuals with type I- or type III-specific reactions. Well-characterized reference sera and further specific peptide markers are needed to establish and to perform future serotyping approaches with higher resolution.     

Sarcoidosis and tuberculosis cytokine profiles: indistinguishable in bronchoalveolar lavage but different in blood

Background

The clinical, radiological and pathological similarities between sarcoidosis and tuberculosis can make disease differentiation challenging. A complicating factor is that some cases of sarcoidosis may be initiated by mycobacteria. We hypothesised that immunological profiling might provide insight into a possible relationship between the diseases or allow us to distinguish between them.

Methods

We analysed bronchoalveolar lavage (BAL) fluid in sarcoidosis (n = 18), tuberculosis (n = 12) and healthy volunteers (n = 16). We further investigated serum samples in the same groups; sarcoidosis (n = 40), tuberculosis (n = 15) and healthy volunteers (n = 40). A cross-sectional analysis of multiple cytokine profiles was performed and data used to discriminate between samples.

Results

We found that BAL profiles were indistinguishable between both diseases and significantly different from healthy volunteers. In sera, tuberculosis patients had significantly lower levels of the Th2 cytokine interleukin-4 (IL-4) than those with sarcoidosis (p = 0.004). Additional serum differences allowed us to create a linear regression model for disease differentiation (within-sample accuracy 91%, cross-validation accuracy 73%).

Conclusions

These data warrant replication in independent cohorts to further develop and validate a serum cytokine signature that may be able to distinguish sarcoidosis from tuberculosis. Systemic Th2 cytokine differences between sarcoidosis and tuberculosis may also underly different disease outcomes to similar respiratory stimuli.     

Epidemiologic behavior of obesity in the Maracaibo City metabolic syndrome prevalence study

Introduction

Obesity is a worldwide public health issue. Since the epidemiological behaviour of this disease is not well established in our country, the purpose of this study was to determinate its prevalence in the Maracaibo City, Zulia State- Venezuela.

Materials and Methods

A cross-sectional study was undertaken using the data set from the Maracaibo City Metabolic Syndrome Prevalence Study. The sample consists of 2108 individuals from both genders and randomly selected: 1119 (53.09%) women and 989 (46.91%) men. The participants were interrogated for a complete clinical history and anthropometric measurements. To classify obesity, the WHO criteria for Body Mass Index (BMI), and Waist Circumference (WC) from the IDF/NHLBI/AHA/WHF/IAS/IASO-2009 (IDF-2009) and ATPIII statements were applied.

Results

For BMI, obesity had an overall prevalence of 33.3% (n = 701), and according to gender women had 32.4% (n = 363) and men had 34.2% (n = 338). Overweight had a prevalence of 34.8% (n = 733), Normal weight had 29.8% (n = 629), and Underweight had 2.1% (n = 45). Adding Obesity and Overweight results, the prevalence of elevated BMI (>25 Kg/m²) was 68.1%. Using the IDF-2009 WC''s cut-off, Obesity had 74.2% prevalence, compared to 51.7% using the ATPIII parameters.

Conclusions

These results show a high prevalence of abdominal obesity in our locality defined by the WHO, IDF-2009 and ATPIII criteria, which were not designed for Latin-American populations. We suggest further investigation to estimate the proper values according to ethnicity, genetic background and sociocultural aspects.     

Genetic association study identifies HSPB7 as a risk gene for idiopathic dilated cardiomyopathy

Dilated cardiomyopathy (DCM) is a structural heart disease with strong genetic background. Monogenic forms of DCM are observed in families with mutations located mostly in genes encoding structural and sarcomeric proteins. However, strong evidence suggests that genetic factors also affect the susceptibility to idiopathic DCM. To identify risk alleles for non-familial forms of DCM, we carried out a case-control association study, genotyping 664 DCM cases and 1,874 population-based healthy controls from Germany using a 50K human cardiovascular disease bead chip covering more than 2,000 genes pre-selected for cardiovascular relevance. After quality control, 30,920 single nucleotide polymorphisms (SNP) were tested for association with the disease by logistic regression adjusted for gender, and results were genomic-control corrected. The analysis revealed a significant association between a SNP in HSPB7 gene (rs1739843, minor allele frequency 39%) and idiopathic DCM (p = 1.06×10⁻⁶, OR = 0.67 [95% CI 0.57–0.79] for the minor allele T). Three more SNPs showed p < 2.21×10⁻⁵. De novo genotyping of these four SNPs was done in three independent case-control studies of idiopathic DCM. Association between SNP rs1739843 and DCM was significant in all replication samples: Germany (n = 564, n = 981 controls, p = 2.07×10⁻³, OR = 0.79 [95% CI 0.67–0.92]), France 1 (n = 433 cases, n = 395 controls, p = 3.73×10⁻³, OR = 0.74 [95% CI 0.60–0.91]), and France 2 (n = 249 cases, n = 380 controls, p = 2.26×10⁻⁴, OR = 0.63 [95% CI 0.50–0.81]). The combined analysis of all four studies including a total of n = 1,910 cases and n = 3,630 controls showed highly significant evidence for association between rs1739843 and idiopathic DCM (p = 5.28×10⁻¹³, OR = 0.72 [95% CI 0.65–0.78]). None of the other three SNPs showed significant results in the replication stage.This finding of the HSPB7 gene from a genetic search for idiopathic DCM using a large SNP panel underscores the influence of common polymorphisms on DCM susceptibility.     

The Guinea-Bissau family of Mycobacterium tuberculosis complex revisited

The Guinea-Bissau family of strains is a unique group of the Mycobacterium tuberculosis complex that, although genotypically closely related, phenotypically demonstrates considerable heterogeneity. We have investigated 414 M. tuberculosis complex strains collected in Guinea-Bissau between 1989 and 2008 in order to further characterize the Guinea-Bissau family of strains. To determine the strain lineages present in the study sample, binary outcomes of spoligotyping were compared with spoligotypes existing in the international database SITVIT2. The major circulating M. tuberculosis clades ranked in the following order: AFRI (n = 195, 47.10%), Latin-American-Mediterranean (LAM) (n = 75, 18.12%), ill-defined T clade (n = 53, 12.8%), Haarlem (n = 37, 8.85%), East-African-Indian (EAI) (n = 25, 6.04%), Unknown (n = 12, 2.87%), Beijing (n = 7, 1.68%), X clade (n = 4, 0.96%), Manu (n = 4, 0.97%), CAS (n = 2, 0.48%). Two strains of the LAM clade isolated in 2007 belonged to the Cameroon family (SIT61). All AFRI isolates except one belonged to the Guinea-Bissau family, i.e. they have an AFRI_1 spoligotype pattern, they have a distinct RFLP pattern with low numbers of IS6110 insertions, and they lack the regions of difference RD7, RD8, RD9 and RD10, RD701 and RD702. This profile classifies the Guinea-Bissau family, irrespective of phenotypic biovar, as part of the M. africanum West African 2 lineage, or the AFRI_1 sublineage according to the spoligtyping nomenclature. Guinea-Bissau family strains display a variation of biochemical traits classically used to differentiate M. tuberculosis from M. bovis. Yet, the differential expression of these biochemical traits was not related to any genes so far investigated (narGHJI and pncA). Guinea-Bissau has the highest prevalence of M. africanum recorded in the African continent, and the Guinea-Bissau family shows a high phylogeographical specificity for Western Africa, with Guinea-Bissau being the epicenter. Trends over time however indicate that this family of strains is waning in most parts of Western Africa, including Guinea-Bissau (p = 0.048).     

Bone related health status in adolescent cyclists

Purpose

To describe bone status and analyse bone mass in adolescent cyclists.

Methods

Male road cyclists (n = 22) who had been training for a minimum of 2 years and a maximum of 7 years with a volume of 10 h/w, were compared to age-matched controls (n = 22) involved in recreational sports activities. Subjects were divided in 2 groups based on age: adolescents under 17 yrs (cyclists, n = 11; controls, n = 13) and over 17 yrs (cyclists, n = 11; controls, n = 9). Peak oxygen uptake (VO₂max) was measured on a cycloergometer. Whole body, lumbar spine, and hip bone mineral content (BMC), density (BMD) and bone area were assessed using dual x-ray absorptiometry (DXA). Volumetric BMD (vBMD) and bone mineral apparent density (BMAD) were also estimated.

Results

The BMC of cyclists was lower for the whole body, pelvis, femoral neck and legs; BMD for the pelvis, hip, legs and whole body and legs bone area was lower but higher in the hip area (all, P≤0.05) after adjusting by lean mass and height. The BMC of young cyclists was 10% lower in the leg and 8% higher in the hip area than young controls (P≤0.05). The BMC of cyclists over 17 yrs was 26.5%, 15.8% and 14.4% lower BMC at the pelvis, femoral neck and legs respectively while the BMD was 8.9% to 24.5% lower for the whole body, pelvis, total hip, trochanter, intertrochanter, femoral neck and legs and 17.1% lower the vBMD at the femoral neck (all P≤0.05). Grouped by age interaction was found in both pelvis and hip BMC and BMD and in femoral neck vBMD (all P≤0.05).

Conclusion

Cycling performed throughout adolescence may negatively affect bone health, then compromising the acquisition of peak bone mass.     

A preliminary evaluation of limb salvage surgery for osteosarcoma around knee joint

Objective

To evaluate the effectiveness and drawbacks of diversified procedures of limb salvage surgery (LSS), providing a reference of rational surgical criterion of LSS.

Methods

Fifty eight patients with stage IIB extremity osteosarcoma around knee joint area between 1992 and 2002 were studied retrospectively. Among them, 43 patients were treated by LSS followed by reconstruction. Reconstruction approaches included re-implantation of irradiation-devitalized tumor bone (n = 12), autoclaving-devitalized tumor bone (n = 8), prosthetic replacement (n = 11), allograft transplantation (n = 8) and vascularized fibula autograft implantation (n = 4). Amputations were performed in 15 patients. Patients were followed up for 6–16 years.

Results

There were no significant difference between LSS and amputation groups regarding disease free survival and local recurrence rates. The actuarial 5-year continuous disease free survival and local recurrence rate were 30.0% and 25.0% in patients of devitalized LSS group, whereas those were 56.5% and 8.7% in patients of non-devitalized reconstruction group. The complication rate was significantly higher in LSS group compared to amputation group (P = 0.003).

Conclusion

LSS with non-devitalized procedures is the optimal treatment for osteosarcoma around knee joint area. Prosthesis implantation is the preferred option for bone reconstruction following LSS. Prevention and treatment of post-operative complications should be paid more attention to get good long-term outcomes of surgery.     

The dynamics of naturally acquired immune responses to Plasmodium falciparum sexual stage antigens Pfs230 & Pfs48/45 in a low endemic area in Tanzania

Background

Naturally acquired immune responses against sexual stages of P. falciparum can reduce the transmission of malaria from humans to mosquitoes. These antigens are candidate transmission-blocking vaccines but little is known about the acquisition of sexual stage immunity after exposure to gametocytes, or their longevity and functionality. We conducted a longitudinal study on functional sexual stage immune responses.

Methodology/Principal Findings

Parasitaemic individuals (n = 116) were recruited at a health centre in Lower Moshi, Tanzania. Patients presented with gametocytes (n = 16), developed circulating gametocytes by day 7 (n = 69) or between day 7 and 14 (n = 10) after treatment or did not develop gametocytes (n = 21). Serum samples were collected on the first day of gametocytaemia and 28 and 84 days post-enrolment (or d7, 28, 84 after enrolment from gametocyte-negative individuals). Antibody responses to sexual stage antigens Pfs230 and Pfs48/45 were detected in 20.7% (72/348) and 15.2% (53/348) of the samples, respectively, and were less prevalent than antibodies against asexual stage antigens MSP-1₁₉ (48.1%; 137/285) and AMA-1 (52.4%; 129/246)(p<0.001). The prevalence of anti-Pfs230 (p = 0.026) and anti-Pfs48/45 antibodies (p = 0.017) increased with longer duration of gametocyte exposure and had an estimated half-life of approximately 3 months. Membrane feeding experiments demonstrated a strong association between the prevalence and concentration of Pfs230 and Pfs48/45 antibodies and transmission reducing activity (TRA, p<0.01).

Conclusions/Significance

In a longitudinal study, anti-Pfs230 and Pf48/45 antibodies developed rapidly after exposure to gametocytes and were strongly associated with transmission-reducing activity. Our data indicate that the extent of antigen exposure is important in eliciting functional transmission-reducing immune responses.     

Clinical characteristics of 26 human cases of highly pathogenic avian influenza A (H5N1) virus infection in China

Background

While human cases of highly pathogenic avian influenza A (H5N1) virus infection continue to increase globally, available clinical data on H5N1 cases are limited. We conducted a retrospective study of 26 confirmed human H5N1 cases identified through surveillance in China from October 2005 through April 2008.

Methodology/Principal Findings

Data were collected from hospital medical records of H5N1 cases and analyzed. The median age was 29 years (range 6–62) and 58% were female. Many H5N1 cases reported fever (92%) and cough (58%) at illness onset, and had lower respiratory findings of tachypnea and dyspnea at admission. All cases progressed rapidly to bilateral pneumonia. Clinical complications included acute respiratory distress syndrome (ARDS, 81%), cardiac failure (50%), elevated aminotransaminases (43%), and renal dysfunction (17%). Fatal cases had a lower median nadir platelet count (64.5×10⁹ cells/L vs 93.0×10⁹ cells/L, p = 0.02), higher median peak lactic dehydrogenase (LDH) level (1982.5 U/L vs 1230.0 U/L, p = 0.001), higher percentage of ARDS (94% [n = 16] vs 56% [n = 5], p = 0.034) and more frequent cardiac failure (71% [n = 12] vs 11% [n = 1], p = 0.011) than nonfatal cases. A higher proportion of patients who received antiviral drugs survived compared to untreated (67% [8/12] vs 7% [1/14], p = 0.003).

Conclusions/Significance

The clinical course of Chinese H5N1 cases is characterized by fever and cough initially, with rapid progression to lower respiratory disease. Decreased platelet count, elevated LDH level, ARDS and cardiac failure were associated with fatal outcomes. Clinical management of H5N1 cases should be standardized in China to include early antiviral treatment for suspected H5N1 cases.     

MRI discriminates thrombus composition and ST resolution after percutaneous coronary intervention in patients with ST-elevation myocardial infarction

Histological composition of material obtained by thrombus aspiration during percutaneous coronary intervention (PCI) in patients with ST-segment elevation acute myocardial infarction (STEMI) is highly variable. We aimed to characterize this material using magnetic resonance imaging (MRI) and to correlate MRI findings with the success of PCI in terms of ST-segment resolution. Thrombus aspiration during primary or rescue PCI was attempted in 100 consecutive STEMI patients, of whom enough material for MRI was obtained in 59. MR images were obtained at 9.4T and T1 and T2 values were measured. Patients with (n = 31) and without (n = 28) adequate ST resolution 120 min after PCI (≥70% of pre-PCI value) had similar baseline characteristics except for a higher prevalence of diabetes mellitus in the latter (10 vs. 43%, p = 0.003). T1 values were similar in both groups (1248±112 vs. 1307±85 ms, respectively, p = 0.7). T2 values averaged 31.2±10.3 and 36.6±12.2 ms; in thrombus from patients with and without adequate ST resolution (p = 0.09). After adjusting for diabetes and other baseline characteristics, lower T2 values were significantly associated with inadequate ST resolution (odds ratio for 1 ms increase 1.08, CI 95% 1.01–1.16, p = 0.027). Histology classified thrombus in 3 groups: coagulated blood (n = 38), fibrin rich (n = 9) and lipid-rich (n = 3). Thrombi composed mostly of coagulated blood were characterized as being of short (n = 10), intermediate (n = 15) or long evolution (n = 13), T2 values being 34.0±13.2, 31.9±8.3 and 31.5±7.9 ms respectively (p = NS). In this subgroup, T2 was significantly higher in specimens from patients with inadequate perfusion (35.9±10.3 versus 28.6±6.7 ms, p = 0.02). This can be of clinical interest as it provides information on the probability of adequate ST resolution, a surrogate for effective myocardial reperfusion.     

Effects of preoperative aspirin on cardiocerebral and renal complications in non-emergent cardiac surgery patients: a sub-group and cohort study

Background and Objective

Postoperative cardiocerebral and renal complications are a major threat for patients undergoing cardiac surgery. This study was aimed to examine the effect of preoperative aspirin use on patients undergoing cardiac surgery.

Methods

An observational cohort study was performed on consecutive patients (n = 1879) receiving cardiac surgery at this institution. The patients excluded from the study were those with preoperative anticoagulants, unknown aspirin use, or underwent emergent cardiac surgery. Outcome events included were 30-day mortality, renal failure, readmission and a composite outcome - major adverse cardiocerebral events (MACE) that include permanent or transient stroke, coma, perioperative myocardial infarction (MI), heart block and cardiac arrest.

Results

Of all patients, 1145 patients met the inclusion criteria and were divided into two groups: those taking (n = 858) or not taking (n = 287) aspirin within 5 days preceding surgery. Patients with aspirin presented significantly more with history of hypertension, diabetes, peripheral arterial disease, previous MI, angina and older age. With propensity scores adjusted and multivariate logistic regression, however, this study showed that preoperative aspirin therapy (vs. no aspirin) significantly reduced the risk of MACE (8.4% vs. 12.5%, odds ratio [OR] 0.585, 95% CI 0.355–0.964, P = 0.035), postoperative renal failure (2.6% vs. 5.2%, OR 0.438, CI 0.203–0.945, P = 0.035) and dialysis required (0.8% vs. 3.1%, OR 0.230, CI 0.071–0.742, P = 0.014), but did not significantly reduce 30-day mortality (4.1% vs. 5.8%, OR 0.744, CI 0.376–1.472, P = 0.396) nor it increased readmissions in the patients undergoing cardiac surgery.

Conclusions

Preoperative aspirin therapy is associated with a significant decrease in the risk of MACE and renal failure and did not increase readmissions in patients undergoing non-emergent cardiac surgery.     

Homogeneous datasets of triple negative breast cancers enable the identification of novel prognostic and predictive signatures

Background

Current prognostic gene signatures for breast cancer mainly reflect proliferation status and have limited value in triple-negative (TNBC) cancers. The identification of prognostic signatures from TNBC cohorts was limited in the past due to small sample sizes.

Methodology/Principal Findings

We assembled all currently publically available TNBC gene expression datasets generated on Affymetrix gene chips. Inter-laboratory variation was minimized by filtering methods for both samples and genes. Supervised analysis was performed to identify prognostic signatures from 394 cases which were subsequently tested on an independent validation cohort (n = 261 cases).

Conclusions/Significance

Using two distinct false discovery rate thresholds, 25% and <3.5%, a larger (n = 264 probesets) and a smaller (n = 26 probesets) prognostic gene sets were identified and used as prognostic predictors. Most of these genes were positively associated with poor prognosis and correlated to metagenes for inflammation and angiogenesis. No correlation to other previously published prognostic signatures (recurrence score, genomic grade index, 70-gene signature, wound response signature, 7-gene immune response module, stroma derived prognostic predictor, and a medullary like signature) was observed. In multivariate analyses in the validation cohort the two signatures showed hazard ratios of 4.03 (95% confidence interval [CI] 1.71–9.48; P = 0.001) and 4.08 (95% CI 1.79–9.28; P = 0.001), respectively. The 10-year event-free survival was 70% for the good risk and 20% for the high risk group. The 26-gene signatures had modest predictive value (AUC = 0.588) to predict response to neoadjuvant chemotherapy, however, the combination of a B-cell metagene with the prognostic signatures increased its response predictive value. We identified a 264-gene prognostic signature for TNBC which is unrelated to previously known prognostic signatures.     

Aberrant hippocampal subregion networks associated with the classifications of aMCI subjects: a longitudinal resting-state study

Background

Altered hippocampal structure and function is a valuable indicator of possible conversion from amnestic type mild cognitive impairment (aMCI) to Alzheimer''s disease (AD). However, little is known about the disrupted functional connectivity of hippocampus subregional networks in aMCI subjects.

Methodology/Principal Findings

aMCI group-1 (n = 26) and controls group-1 (n = 18) underwent baseline and after approximately 20 months follow up resting-state fMRI scans. Integrity of distributed functional connectivity networks incorporating six hippocampal subregions (i.e. cornu ammonis, dentate gyrus and subicular complex, bilaterally) was then explored over time and comparisons made between groups. The ability of these extent longitudinal changes to separate unrelated groups of 30 subjects (aMCI-converters, n = 6; aMCI group-2, n = 12; controls group-2, n = 12) were further assessed. Six longitudinal hippocampus subregional functional connectivity networks showed similar changes in aMCI subjects over time, which were mainly associated with medial frontal gyrus, lateral temporal cortex, insula, posterior cingulate cortex (PCC) and cerebellum. However, the disconnection of hippocampal subregions and PCC may be a key factor of impaired episodic memory in aMCI, and the functional index of these longitudinal changes allowed well classifying independent samples of aMCI converters from non-converters (sensitivity was 83.3%, specificity was 83.3%) and controls (sensitivity was 83.3%, specificity was 91.7%).

Conclusions/Significance

It demonstrated that the functional changes in resting-state hippocampus subregional networks could be an important and early indicator for dysfunction that may be particularly relevant to early stage changes and progression of aMCI subjects.     

Antiretroviral therapy outcomes in HIV-infected children after adjusting protease inhibitor dosing during tuberculosis treatment

Background

Modification of ritonavir-boosted lopinavir (LPV/r)-based antiretroviral therapy is required for HIV-infected children co-treated for tuberculosis (TB). We aimed to determine virologic and toxicity outcomes among TB/HIV co-treated children with the following modifications to their antiretroviral therapy (ART): (1) super-boosted LPV/r, (2) double-dose LPV/r or (3) ritonavir.

Methods and Findings

A medical record review was conducted at two clinical sites in Johannesburg, South Africa. The records of children 6–24 months of age initiating LPV/r-based therapy were reviewed. Children co-treated for TB were categorized based on the modifications made to their ART regimen and were compared to children of the same age at each site not treated for TB.Included are 526 children, 294 (56%) co-treated for TB. All co-treated children had more severe HIV disease, including lower CD4 percents and worse growth indicators, than comparisons.Children in the super-boosted group (n = 156) were as likely to be virally suppressed (<400 copies/ml) at 6 months as comparisons (69.2% vs. 74.8%, p = 0.36). Children in the double-dose (n = 47) and ritonavir groups (n = 91) were significantly less likely to be virally suppressed at 6 months (53.1% and 49.3%) than comparisons (74.8% and 82.1%; p = 0.02 and p<0.0001, respectively). At 12 months only children in the ritonavir group still had lower rates of virological suppression relative to comparisons (63.9% vs 83.3% p<0.05). Grade 1 or greater ALT elevations were more common in the super-boosted (75%) than double-dose (54.6%) or ritonavir (33.9%) groups (p = 0.09 and p<0.0001) but grade 3/4 elevations were observed in 3 (13.6%) of the super-boosted, 7 (15.9%) of the double-dose and 5 (8.9%) of the ritonavir group (p = 0.81 and p = 0.29).

Conclusion

Good short-term virologic outcomes were achieved in children co-treated for TB and HIV who received super-boosted LPV/r. Treatment limiting toxicity was rare. Strategies for increased dosing of LPV/r with TB treatment warrant further investigation.     

Efficient removal of platelets from peripheral blood progenitor cell products using a novel micro-chip based acoustophoretic platform

Background

Excessive collection of platelets is an unwanted side effect in current centrifugation-based peripheral blood progenitor cell (PBPC) apheresis. We investigated a novel microchip-based acoustophoresis technique, utilizing ultrasonic standing wave forces for the removal of platelets from PBPC products. By applying an acoustic standing wave field onto a continuously flowing cell suspension in a micro channel, cells can be separated from the surrounding media depending on their physical properties.

Study Design and Methods

PBPC samples were obtained from patients (n = 15) and healthy donors (n = 6) and sorted on an acoustophoresis-chip. The acoustic force was set to separate leukocytes from platelets into a target fraction and a waste fraction, respectively. The PBPC samples, the target and the waste fractions were analysed for cell recovery, purity and functionality.

Results

The median separation efficiency of leukocytes to the target fraction was 98% whereas platelets were effectively depleted by 89%. PBPC samples and corresponding target fractions were similar in the percentage of CD34+ hematopoetic progenitor/stem cells as well as leukocyte/lymphocyte subset distributions. Median viability was 98%, 98% and 97% in the PBPC samples, the target and the waste fractions, respectively. Results from hematopoietic progenitor cell assays indicated a preserved colony-forming ability post-sorting. Evaluation of platelet activation by P-selectin (CD62P) expression revealed a significant increase of CD62P+ platelets in the target (19%) and waste fractions (20%), respectively, compared to the PBPC input samples (9%). However, activation was lower when compared to stored blood bank platelet concentrates (48%).

Conclusion

Acoustophoresis can be utilized to efficiently deplete PBPC samples of platelets, whilst preserving the target stem/progenitor cell and leukocyte cell populations, cell viability and progenitor cell colony-forming ability. Acoustophoresis is, thus, an interesting technology to improve current cell processing methods.     

Mycobacterium tuberculosis Population in Northwestern Russia: An Update from Russian-EU/Latvian Border Region

This study aimed to characterize the population structure of Mycobacterium tuberculosis in Pskov oblast in northwestern Russia, to view it in the geographical context, to compare drug resistance properties across major genetic families. Ninety M. tuberculosis strains from tuberculosis (TB) patients, permanent residents in Pskov oblast were subjected to LAM-specific IS6110-PCR and spoligotyping, followed by comparison with SITVITWEB and MIRU-VNTRplus databases. The Beijing genotype (n = 40) was found the most prevalent followed by LAM (n = 18), T (n = 13), Haarlem (n = 10), Ural (n = 5), and Manu2 (n = 1); the family status remained unknown for 3 isolates. The high rate of Beijing genotype and prevalence of LAM family are similar to those in the other Russian settings. A feature specific for M. tuberculosis population in Pskov is a relatively higher rate of Haarlem and T types. Beijing strains were further typed with 12-MIRU (followed by comparison with proprietary global database) and 3 hypervariable loci QUB-3232, VNTR-3820, VNTR-4120. The 12-MIRU typing differentiated 40 Beijing strains into 14 types (HGI = 0.82) while two largest types were M2 (223325153533) prevalent throughout former USSR and M11 (223325173533) prevalent in Russia and East Asia. The use of 3 hypervariable loci increased a discrimination of the Beijing strains (18 profiles, HGI = 0.89). Both major families Beijing and LAM had similar rate of MDR strains (62.5 and 55.6%, respectively) that was significantly higher than in other strains (21.9%; P = 0.001 and 0.03, respectively). The rpoB531 mutations were more frequently found in Beijing strains while LAM drug resistant strains mainly harbored rpoB516 and inhA −15 mutations. Taken together with a high rate of multidrug resistance among Beijing strains from new TB cases (79.3% versus 44.4% in LAM), these findings suggest the critical impact of the Beijing genotype on the current situation with MDR-TB in the Pskov region in northwestern Russia.