Male spondyloarthritis patients and those with longer disease duration have less severe disc degeneration: propensity score-matched comparison.

Objective: Using whole spine sagittal T2 MRI, we aimed to compare the severity and prevalence of disc degeneration (DD) in axial SpA patients vs the general population and to determine any association between spinal inflammation, structural changes, mobility and DD among SpA patients. Methods: Two prospectively collected cohorts of SpA patients (n = 411) and the general population (n = 2007) were recruited. Eventually, 967 participants from the populational cohort and 304 participants from the SpA cohort were analysed. Two hundred and nineteen matched pairs were generated by propensity score matching. Imaging parameters, including Pfirrmann grading, disc herniation, high-intensity zone, Schmorl's node, Modic change and anterior marrow change were studied and compared from C2/3 to L5/S1. DD was defined as Pfirrmann grade 4 or 5. Demographic factors, including age, sex and BMI, were collected. Multivariable linear regression was used to determine the association between spinal inflammation [Spondyloarthritis Research Consortium of Canada (SPARCC) spine MRI index], structural changes [modified Stoke Ankylosing Spondylitis Spinal Score (mSASSS)] and mobility (BASMI) with lumbar Pfirrmann score. Results: SpA patients had lower prevalence of DD (P < 0.001). The disease stage-stratified regression model showed that SPARCC spinal MRI index was associated with higher lumbar Pfirrmann scores in early disease (ß = 0.196, P = 0.044), whereas mSASSS was associated with lower lumbar Pfirrmann scores in later disease (ß = -0.138, P = 0.038). Males had higher mSASSS (P < 0.001) and lower odds of whole spine DD (odds ratio = 0.622, P = 0.028). Conclusion: SpA patients had lower DD severity than the general population. Males had higher mSASSSs, and increased mSASSS at later disease was associated with less severe DD.

Why Are Some Intervertebral Discs More Prone to Degeneration?: Insights Into Isolated Thoracic "Dysgeneration".

STUDY DESIGN: Prospective observational study. OBJECTIVE: To determine the prevalence of isolated thoracic degeneration on magnetic resonance imaging (MRI), demographic factors and imaging features, as well as the patient-reported quality of life outcomes associated with this condition. SUMMARY OF BACKGROUND DATA: Thoracic intervertebral discs are least susceptible to disc degeneration (DD) and may represent a manifestation of "dysgeneration." These discs may never be hydrated from the beginning and seem hypointense on MRI. PATIENTS AND METHODS: A population-based MRI study of 2007 volunteers was conducted. Each disc from C2/3 to L5/S1 was measured by Pfirrmann and Schneiderman grading. Disc herniation, Schmorl node (SN), high-intensity zones (HIZ), and Modic changes were studied. DD was defined by Pfirrmann 4 or 5. patient-reported quality of life scores, including a 36-item short-form questionnaire and visual analog scale for low back pain, were recorded. Subjects were divided into "isolated thoracic degeneration" (only thoracic segment) and "tandem thoracic degeneration" (thoracic with other segments). The association between imaging findings and isolated thoracic degeneration was determined using multivariate logistic regression. RESULTS: The mean age of the subjects was 50.0 ± 0.5 and 61.4% were females (n = 1232). Isolated thoracic degeneration was identified in 2.3% of the cohort. Factors associated with isolated thoracic degeneration included lower age, C6/7 HIZ, T8/9 HIZ, and T8/9 SN. Factors associated with tandem thoracic degeneration included L4/5 posterior bulging. The thoracic and lumbar tandem degeneration group demonstrated higher bodily pain, despite a lower visual analog scale, and a higher physical component score of the 36-item short form. CONCLUSIONS: Isolated thoracic degeneration demonstrated an earlier age of onset, mostly involving the mid-thoracic region (T5/6-T8/9), and in association with findings such as SN. Subjects with tandem thoracolumbar degeneration had less severe lumbar DD and low back pain as compared with those with isolated lumbar degeneration. This paints the picture of "dysgeneration" occurring in the thoracic and lumbar spine. LEVEL OF EVIDENCE: 1.

Multiplexed Phagosomal Assays for the Detection and Quantification of Bidirectional Exchange Between the Phagolysosomal Lumen and Extracellular Space.

The phagolysosome is an antimicrobial and degradative organelle that plays a key role in macrophage-mediated inflammation and homeostasis. Before being presented to the adaptive immune system, phagocytosed proteins must first be processed into immunostimulatory antigens. Until recently, little attention has been given to how other processed PAMPs and DAMPs can stimulate an immune response if they are sequestered in the phagolysosome. Eructophagy is a newly described process in macrophages that releases partially digested immunostimulatory PAMPs and DAMPs extracellularly from the mature phagolysosome to activate vicinal leukocytes. This chapter outlines approaches to observe and quantify eructophagy by simultaneously measuring several phagosomal parameters of individual phagosomes. These methods use specifically designed experimental particles capable of conjugating to multiple reporter/reference fluors in combination with real-time automated fluorescent microscopy. Through the use of high-content image analysis software, each phagosomal parameter can be evaluated quantitatively or semiquantitatively during post-analysis.

Macrophages disseminate pathogen associated molecular patterns through the direct extracellular release of the soluble content of their phagolysosomes.

Recognition of pathogen-or-damage-associated molecular patterns is critical to inflammation. However, most pathogen-or-damage-associated molecular patterns exist within intact microbes/cells and are typically part of non-diffusible, stable macromolecules that are not optimally immunostimulatory or available for immune detection. Partial digestion of microbes/cells following phagocytosis potentially generates new diffusible pathogen-or-damage-associated molecular patterns, however, our current understanding of phagosomal biology would have these molecules sequestered and destroyed within phagolysosomes. Here, we show the controlled release of partially-digested, soluble material from phagolysosomes of macrophages through transient, iterative fusion-fission events between mature phagolysosomes and the plasma membrane, a process we term eructophagy. Eructophagy is most active in proinflammatory macrophages and further induced by toll like receptor engagement. Eructophagy is mediated by genes encoding proteins required for autophagy and can activate vicinal cells by release of phagolysosomally-processed, partially-digested pathogen associated molecular patterns. We propose that eructophagy allows macrophages to amplify local inflammation through the processing and dissemination of pathogen-or-damage-associated molecular patterns.

The impact of short-term fenestrated scleral lens wear on intraocular pressure.

PURPOSE: To examine the influence of short-term fenestrated scleral lens wear on intraocular pressure (IOP) in healthy eyes. METHODS: IOP was measured before, during, and after a brief period (1-2 min) of fenestrated, scleral lens wear, using a rebound tonometer, in fifty, young healthy adults (mean age 23 ± 4 years) with normal corneas. RESULTS: Immediately following lens insertion, 48 of the 50 (96 %) of participants displayed an increase in IOP (mean ± SD increase in these participants of 3.8 ± 2.0 mmHg). Immediately following lens removal, 50 % of participants displayed a reduction in IOP, equal to or lower than, the pre-insertion IOP (-1.0 ± 0.8 mmHg lower than baseline). The remaining 50 % of participants displayed an IOP slightly greater than the pre-insertion IOP (1.6 ± 1.0 mmHg greater) after lens removal. CONCLUSIONS: Short-term fenestrated scleral lens wear resulted in a small, but statistically significant, increase in IOP in 96 % of young healthy participants (< 4 mmHg on average), which decreased to a level similar to pre-lens insertion IOP levels immediately following lens removal. Further research is required to determine if this measured change in IOP during scleral lens wear is artefactual, or an elevation in the true IOP.

Simultaneous Analysis of Multiple Lumenal Parameters of Individual Phagosomes Using High-Content Imaging.

The biochemical processes within the phagosomes of macrophages and dendritic cells are essential to immunity and homeostasis. Measurable properties of the phagosomal lumen include: assessment of various hydrolytic activities, reduction and oxidation events, pH, ion concentrations, and electrochemical gradients. These often-interdependent phagosomal features are commonly evaluated individually, hindering the analysis of the biochemical relationship between these factors within the same phagosome. In addition, the ability of phagosomes within the same cell to behave autonomously is becoming more evident, thus highlighting the need for a technique capable of multiplex analyses of phagosomal lumenal chemistries at the single phagosome level. In this chapter, we outline an approach that is capable of simultaneously measuring multiple phagosomal parameters of individual phagosomes by utilizing specifically designed experimental particles with multiple reporter fluors, in combination with real-time fluorometric measurement via automated microscopy. Subsequent analysis using high-content image analysis software enables each phagosomal parameter to be evaluated in a quantitative or semi-quantitative manner. This approach facilitates investigation of the complex relationship between phagosomal properties in a population of macrophages in real time, at the level of individual phagosomes.

Pyk2 Controls Integrin-Dependent CTL Migration through Regulation of De-Adhesion.

Protein tyrosine kinase 2 (Pyk2) is required for T cell adhesion to ICAM-1; however, the mechanism by which it regulates adhesion remains unexplored. Pyk2 function in murine CTL clones and activated ex vivo CD8(+) T cells was disrupted by pharmacological inhibition, knockdown of expression with small interfering RNA, or expression of the dominant-negative C-terminal domain. We found that Pyk2 is not absolutely required for adhesion of CTL to ICAM-1, but rather delays the initial adhesion. Disruption of Pyk2 function caused cells to display an unusual elongated appearance after 1 h on ICAM-1, consistent with abnormally strong adhesion. Furthermore, the random mobility of CTL on ICAM-1 was severely compromised using all three methods of disrupting Pyk2 function. Live-cell imaging studies revealed that the decreased migration is the result of a defect in the detachment from ICAM-1 at the trailing edge when Pyk2 function is inhibited. Examination of Pyk2 tyrosine phosphorylation in normal polarized cells demonstrated that Pyk2 phosphorylated at Y579 and Y580 preferentially localizes to the leading edge, whereas Y881-phosphorylated Pyk2 is enriched at the trailing edge, suggesting that the tyrosine phosphorylation of Pyk2 is spatially regulated in migrating CTL. Additionally, inhibition of Pyk2 caused cells to form multiple LFA-1-rich tails at the trailing edge, most likely resulting from a defect in LFA-1 release required for forward movement. Our results show that Pyk2 contributes to CTL migration by regulating detachment of CTL at the trailing edge, which could explain why Pyk2 is important for chemotactic and migratory responses.

Can pre-operative axial CT imaging predict syndesmosis instability in patients sustaining ankle fractures? Seven years' experience in a tertiary trauma center.

OBJECTIVE: The purpose of this study is to explore the diagnostic accuracy of CT measurements in predicting syndesmosis instability of injured ankle, with correlation to operative findings. METHODS: From July 2006 to June 2013, 123 patients presented to a single tertiary hospital who received pre-operative CT for ankle fractures were retrospectively reviewed. All patients underwent open reduction and internal fixation for fractures and intra-operative syndesmosis integrity tests. The morphology of incisura fibularis was categorized as deep or shallow. The tibiofibular distance (TFD) between the medial border of the fibula and the nearest point of the lateral border of tibia were measured at anterior (aTFD), middle (mTFD), posterior (pTFD), and maximal (maxTFD) portions across the syndesmosis on axial CT images at 10 mm proximal to the tibial plafond. Statistical analysis was performed with independent samples t test and ROC curve analysis. Intraobserver reproducibility and inter-observers agreement were also evaluated. RESULTS: Of the 123 patients, 39 (31.7%) were operatively diagnosed with syndesmosis instability. No significant difference of incisura fibularis morphology (deep or shallow) and TFDs was demonstrated respective to genders. The axial CT measurements were significantly higher in ankles diagnosed with syndesmosis instability than the group without (maxTFD means 7.2 ± 2.96 mm vs. 4.6 ± 1.4 mm, aTFD mean 4.9 ± 3.7 mm vs. 1.8 ± 1.4 mm, mTFD mean 5.3 ± 2.4 mm vs. 3.2 ± 1.6 mm, pTFD mean 5.3 ± 1.8 mm vs. 4.1 ± 1.3 mm, p < 0.05). Their respective cutoff values with best sensitivity and specificity were calculated; the aTFD (AUC 0.798) and maxTFD (AUC 0.794) achieved the highest diagnostic accuracy. The optimal cutoff levels were aTFD = mm (sensitivity, 56.4%; specificity, 91.7%) and maxTFD = 5.65 mm (sensitivity, 74.4%; specificity, 79.8%). The inter-observer agreement was good for all aTFD, mTFD, pTFD, and maxTFD measurements (ICC 0.959, 0.799, 0.783, and 0.865). The ICC for intraobserver agreement was also very good, ranging from 0.826 to 0.923. CONCLUSIONS: Axial CT measurements of tibiofibular distance were useful predictors for syndesmosis instability in fractured ankles. The aTFD and maxTFD are the most powerful parameters to predict positive operative instability.

Development of a candidate certified reference material of cypermethrin in green tea.

This paper presents the preparation of a candidate certified reference material (CRM) of cypermethrin in green tea, GLHK-11-01a according to the requirements of ISO Guide 34 and 35. Certification of the material was performed using a newly developed isotope dilution mass spectrometry (IDMS) approach, with gas chromatography high resolution mass spectrometry (GC-HRMS) and gas chromatography-tandem mass spectrometry (GC-MS/MS). Statistical analysis (one-way ANOVA) showed excellent agreement of the analytical data sets generated from the two mass spectrometric detections. The characterization methods have also been satisfactorily applied in an Asia-Pacific Metrology Program (APMP) interlaboratory comparison study. Both the GC-HRIDMS and GC-IDMS/MS methods proved to be sufficiently reliable and accurate for certification purpose. The certified value of cypermethrin in dry mass fraction was 148 µg kg(-1) and the associated expanded uncertainty was 14µg kg(-1). The uncertainty budget was evaluated from sample in homogeneity, long-term and short-term stability and variability in the characterization procedure. GLHK-11-01a is primarily developed to support the local and wider testing community on need basis in quality assurance work and in seeking accreditation.

The first international proficiency test on ketamine and norketamine in hair.

This paper discusses the organization of the first international proficiency test (PT) programme on ketamine (K) and norketamine (NK) in hair samples. The primary objective of the programme was to evaluate the analytical capability of participating laboratories on hair analysis for K and NK via comparison of results. Authentic samples, instead of spiked samples were used in the programme to mimic the analysis of incorporated illicit drugs in real-life situations. Eight of the ten participating medical or forensic laboratories from Australia, France, Hong Kong, Italy, Singapore and the USA returned results to the organizer. Quantification methods from these laboratories were confined to GC-MS and LC-MS/MS. Performance assessment based on z-score indicated that only three laboratories achieved satisfactory results for both the analysis of K and NK. It was concluded that the overall performance of the participating laboratories was fair and there is still room for further improvement. Additional similarly designed PT programmes are recommended to be organized in order to encourage reliable measurements of illicit drugs in hair samples. Taking into account the substantial effect on the consensus values within limited number of data points, a recommendation on the provision of reference values assigned by accurate methods will be of benefit to small size PT programmes in the forensic field.

Development of melamine certified reference material in milk using two different isotope dilution mass spectrometry techniques.

A new certified reference material (CRM) of melamine in milk GLHK-11-02 was developed aiming to address the great demand from the testing community after the melamine crises. The material was prepared by adding an appropriate quantity of melamine into the skimmed milk samples and the final product was in the form of fine lyophilized powder. Characterization of the material relied on two newly developed gravimetric isotope dilution mass spectrometry (IDMS) methods, one using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and another gas chromatography-mass spectrometry (GC-MS). Experimental parameters with crucial effects on the performance of the two IDMS methods were thoroughly investigated. These included purity of standard used, equilibration time of isotopes, efficiency of extraction methods as well as possible interferences from the matrix and melamine analogues. Precision was found to be excellent with a coefficient of variation of 2.5% for the LC-IDMS/MS (n=46) and 1.9% for the GC-IDMS (n=30) respectively. Using one-tail Student's t-test at 95% confidence interval, analytical data sets generated from the two methods were found to exhibit no significant difference. Measurement accuracy of the methods was further verified through an Asia Pacific Metrology Program (APMP) pilot study. Analytical results of the present LC-IDMS/MS for the two milk test samples at the concentration level of about 0.45 and 3.5 mg kg(-1) were proven to be very good. There were excellent overlaps between our results and the assigned reference values, and the absolute deviation was less than 3.2%. Both the LC-IDMS/MS and GC-IDMS methods were shown to be sufficiently reliable and accurate for certification of the melamine CRM. Certified value of melamine in dry mass fraction in GLHK-11-02 was 1.14 mg kg(-1). Expanded uncertainty due to sample inhomogeneity, long term and short term stability and variability in the characterization procedure was at 7.1% or 0.08 mg kg(-1). The CRM is primarily used to provide a complete method validation for and to improve the technical competence of melamine analysis to food and chemical testing laboratories.

Regulation of the tyrosine kinase Pyk2 by calcium is through production of reactive oxygen species in cytotoxic T lymphocytes.

Pyk2 was identified as a Ca(2+)-dependent kinase, however, the regulation of Pyk2 by Ca(2+) in T cells remains controversial. We found that Ca(2+) mobilization preferentially induced Pyk2 phosphorylation in cytotoxic T lymphocytes (CTL). Furthermore, Pyk2 phosphorylation in CTL was not absolutely Ca(2+) dependent but relied on the strength of T cell receptor stimulation. Ionomycin-stimulated Pyk2 phosphorylation did not require calmodulin activity, because phosphorylation was not inhibited by the calmodulin inhibitor W7, and we detected no Ca(2+)-regulated association between Pyk2 and calmodulin. Ca(2+)-stimulated Pyk2 phosphorylation was dependent on Src-family kinase activity, even at the Pyk2 autophosphorylation site. We sought to identify a Ca(2+)-regulated pathway that could trigger Pyk2 phosphorylation in T cells and found that ionomycin stimulated the production of reactive oxygen species and an H(2)O(2) scavenger inhibited ionomycin-induced Pyk2 phosphorylation. Additionally, H(2)O(2) induced strong Erk activation and ionomycin-stimulated Pyk2 phosphorylation was Erk dependent. These data support the conclusion that Ca(2+) mobilization induces the production of reactive oxygen species, which in turn activate the Erk pathway, leading to Src-family kinase-dependent Pyk2 phosphorylation. Our data demonstrate that Pyk2 is not a Ca(2+)-dependent kinase in T cells but instead, increased intracellular Ca(2+) induces Pyk2 phosphorylation through production of reactive oxygen species. These findings are consistent with the possibility that Pyk2 acts as an early sensor of numerous extracellular signals that trigger a Ca(2+) flux and/or reactive oxygen species to amplify tyrosine phosphorylation signaling events.

Phosphoinositide 3-kinase-regulated adapters in lymphocyte activation.

Signaling via phosphoinositide 3-kinases (PI3Ks) has emerged as a central component of lymphocyte activation via immunoreceptors, costimulatory receptors, cytokine receptors, and chemokine receptors. The discovery of phosphoinositide-binding pleckstrin homology (PH) domains has substantially increased understanding of how PI3Ks activate cellular responses. Accumulating evidence indicates that PH-domain containing adapter molecules provide important links between PI3K and lymphocyte function. Here, we review data on PI3K-regulated adapter proteins of the Grb-associated binder (GAB), Src kinase-associated phosphoprotein (SKAP), and B-lymphocyte adapter molecule of 32 kDa (Bam32)/ dual-adapter for phosphotyrosine and 3-phosphoinositides (DAPP)/TAPP families, with a focus on the latter group. Current data support the model that recruitment of these adapters to the plasma membrane of activated lymphocytes is driven by the phosphoinositides phosphatidylinositol-3,4,5-tris-phosphate and phosphatidylinositol-3,4-bisphosphate, generated through the action of PI3Ks and under the regulatory control of lipid phosphatases Src homology 2 domain-containing inositol phosphatase (SHIP), phosphatase and tensin homolog, and inositol polyphosphate 4-phosphatase. At the plasma membrane, these adapters serve to assemble distinct protein complexes. Bam32/DAPP1 and SKAPs function to promote activation of monomeric guanosine triphosphatases, including Rac and Rap, and promote integrin activation, lymphocyte adhesion to matrix proteins, and cell:cell interactions between B and T lymphocytes. GABs can provide feedforward amplification or feedback inhibition of PI3K signaling. Current work is further defining the molecular interactions driven by these molecules and identifying the functions of TAPP adapters, which also appear to be involved in lymphocyte adhesion and are specific effectors downstream of the SHIP product phosphatidylinositol-3,4-bisphosphate.

TAPP2 links phosphoinositide 3-kinase signaling to B-cell adhesion through interaction with the cytoskeletal protein utrophin: expression of a novel cell adhesion-promoting complex in B-cell leukemia.

Tandem pleckstrin homology domain proteins (TAPPs) are recruited to the plasma membrane via binding to phosphoinositides produced by phosphoinositide 3-kinases (PI3Ks). Whereas PI3Ks are critical for B-cell activation, the functions of TAPP proteins in B cells are unknown. We have identified 40 potential interaction partners of TAPP2 in B cells, including proteins involved in cytoskeletal rearrangement, signal transduction and endocytic trafficking. The association of TAPP2 with the cytoskeletal proteins utrophin and syntrophin was confirmed by Western blotting. We found that TAPP2, syntrophin, and utrophin are coexpressed in normal human B cells and B-chronic lymphocytic leukemia (B-CLL) cells. TAPP2 and syntrophin expression in B-CLL was variable from patient to patient, with significantly higher expression in the more aggressive disease subset identified by zeta-chain-associated protein kinase of 70 kDa (ZAP70) expression and unmutated immunoglobulin heavy chain (IgH) genes. We examined whether TAPP can regulate cell adhesion, a known function of utrophin/syntrophin in other cell types. Expression of membrane-targeted TAPP2 enhanced B-cell adhesion to fibronectin and laminin, whereas PH domain-mutant TAPP2 inhibited adhesion. siRNA knockdown of TAPP2 or utrophin, or treatment with PI3K inhibitors, significantly inhibited adhesion. These findings identify TAPP2 as a novel link between PI3K signaling and the cytoskeleton with potential relevance for leukemia progression.

PI(3,4,5)P3 and PI(3,4)P2 levels correlate with PKB/akt phosphorylation at Thr308 and Ser473, respectively; PI(3,4)P2 levels determine PKB activity.

The PI3K-PKB pathway is an important and widely studied pathway in cell signaling. The enzyme activity of PI3K produces D-3 phosphoinositides, including the lipid second messengers PI(3,4,5)P3 and PI(3,4)P2. PI(3,4,5)P3 has been deemed to be the most important second messenger for triggering PKB phosphorylation. PKB has two regulatory phosphorylation sites, Thr308 and Ser473, both of which contribute to its full activity. The direct relationship between PI3K lipid products and PKB phosphorylation is still not entirely clear. Our previous study showed that PI(3,4)P2 has a specific role in contributing to PKB phosphorylation on Ser473 sites in mast cells. In this study, we used two strategies to further elucidate this question in a well-established B cell system. First, by SHIP overexpression, we examined PKB activation under conditions where PI(3,4,5)P3 accumulation is largely suppressed. Second, we used dose response of different forms of B-cell receptor ligands to manipulate the relative levels of PI(3,4,5)P3 and PI(3,4)P2. Our results demonstrate a close relationship between PI(3,4,5)P3 levels and Thr308 phosphorylation levels, and PI(3,4)P2 levels and Ser473 phosphorylation levels, respectively. Furthermore, overall PKB activity, primarily consisting of cytosolic enzyme, was dependent upon levels of PI(3,4)P2, while only membrane-associated PKB activity was dependent upon PI(3,4,5)P3 levels. We conclude that PI(3,4,5)P3 and PI(3,4)P2 have distinct roles in determining PKB phosphorylation and activity. Thus, when investigating PI3K-PKB pathways, the importance of both lipids must be considered.

Regulation of phosphoinositide 3-kinase signaling by oxidants: hydrogen peroxide selectively enhances immunoreceptor-induced recruitment of phosphatidylinositol (3,4) bisphosphate-binding PH domain proteins.

Phosphoinositide 3-kinases (PI3Ks) generate several distinct lipid second messengers including phosphatidylinositol (3,4,5) trisphosphate (PIP3) and phosphatidylinositol (3,4) bisphosphate PI(3,4)P2. PI(3,4)P2 is produced with distinct kinetics and binds to distinct PH domain effector proteins; however, the regulation of this signaling pathway is poorly understood. Superoxides such as hydrogen peroxide are transiently produced after activation through various cell surface receptors and play important roles in immune and inflammatory responses. Here we use quantitative microscopy to examine the effect of peroxide on PI(3,4)P2-mediated mobilization of signaling proteins in B lymphocytes. Peroxide was found to induce dose-dependant membrane recruitment of the PI(3,4)P2-binding PH domain proteins Bam32, TAPP2 and Akt/PKB but not the PIP3-binding PH domain of Btk. Peroxide-induced membrane recruitment was found to be dependant on PI3K activity, with the p110delta isoform contributing much of the activity in the BJAB human B lymphoma model. Strikingly, peroxide co-stimulation enhanced antigen receptor-induced membrane recruitment of Bam32 and TAPP2, with combined stimulation exceeding the maximum achievable with either stimulus alone. Expression of the lipid phosphatase PTEN led to reduction of antigen receptor-induced membrane recruitment of TAPP2; however, peroxide costimulation could overcome the inhibitory effect of PTEN. Inhibition of the NADPH oxidase led to reduction of antigen receptor-induced membrane recruitment of TAPP2. Our results indicate that exogenous and endogenous superoxides can modulate the quality of the PI3K signal in lymphocytes by selectively increasing PI(3,4)P2-dependant signaling.