A change of heart: Characteristics and outcomes of multiple cardiac retransplant recipients.

BACKGROUND: Among heart transplant (HT) recipients who develop advanced graft dysfunction, cardiac re-transplantation may be considered. A smaller subset of patients will experience failure of their second allograft and undergo repeat re-transplantation. Outcomes among these individuals are not well-described. METHODS: Adult and pediatric patients in the United Network for Organ Sharing (UNOS) registry who received HT between January 1, 1990 and December 31, 2020 were included. RESULTS: Between 1990 and 2020, 90 individuals received a third HT and three underwent a fourth HT. Recipients were younger than those undergoing primary HT (mean age 32 years). Third HT was associated with significantly higher unadjusted rates of 1-year mortality (18% for third HT vs. 13% for second HT vs. 9% for primary HT, p < .001) and 10-year mortality (59% for third HT vs. 42% for second HT vs. 37% for primary HT, p < .001). Mortality was highest amongst recipients aged >60 years and those re-transplanted for acute graft failure. Long-term rates of CAV, rejection, chronic dialysis, and hospitalization for infection were also higher. CONCLUSIONS: Third HT is associated with higher morbidity and mortality than primary HT. Further consensus is needed regarding appropriate organ stewardship for this unique subgroup.

Utility of a fusion protein T-cell co-stimulation blocker Belatacept in heart transplant recipients: Real world experience from a high volume center.

BACKGROUND: Belatacept (BTC), a fusion protein, selectively inhibits T-cell co-stimulation by binding to the CD80 and CD86 receptors on antigen-presenting cells (APCs) and has been used as immunosuppression in adult renal transplant recipients. However, data regarding its use in heart transplant (HT) recipients are limited. This retrospective cohort study aimed to delineate BTC's application in HT, focusing on efficacy, safety, and associated complications at a high-volume HT center. METHODS: A retrospective cohort study was conducted of patients who underwent HT between January 2017 and December 2021 and subsequently received BTC as part of their immunosuppressive regimen. Twenty-one HT recipients were identified. Baseline characteristics, history of rejection, and indication for BTC use were collected. Outcomes included renal function, graft function, allograft rejection and mortality. Follow-up data were collected through December 2023. RESULTS: Among 776 patients monitored from January 2017 to December 2021 21 (2.7%) received BTC treatment. Average age at transplantation was 53 years (± 12 years), and 38% were women. BTC administration began, on average, 689 [483, 1830] days post-HT. The primary indications for BTC were elevated pre-formed donor-specific antibodies in highly sensitized patients (66.6%) and renal sparing (23.8%), in conjunction with reduced calcineurin inhibitor dosage. Only one (4.8%) patient encountered rejection within a year of starting BTC. Graft function by echocardiography remained stable at 6 and 12 months posttreatment. An improvement was observed in serum creatinine levels (76.2% of patients), decreasing from a median of 1.58 to 1.45 (IQR [1.0-2.1] to [1.1-1.9]) over 12 months (p = .054). eGFR improved at 3 and 6 months compared with 3 months pre- BTC levels; however, this was not statistically significant (p = .24). Treatment discontinuation occurred in seven patients (33.3%) of whom four (19%) were switched back to full dose CNI. Infections occurred in 11 patients (52.4%), leading to BTC discontinuation in 4 patients (19%). CONCLUSION: In this cohort, BTC therapy was used as alternative immunosuppression for management of highly sensitized patients or for renal sparing. BTC therapy when combined with CNI dose reduction resulted in stabilization in renal function as measured through renal surrogate markers, which did not, however, reach statistical significance. Patients on BTC maintained a low rejection rate and preserved graft function. Infections were common during BTC therapy and were associated with medication pause/discontinuation in 19% of patients. Further randomized studies are needed to assess the efficacy and safety of BTC in HT recipients.

The utilization of molecular microscope in management of heart transplant recipients in the era of noninvasive monitoring.

INTRODUCTION: Monitoring for graft rejection is a fundamental tenet of post-transplant follow-up. In heart transplantation (HT) in particular, rejection has been traditionally assessed with endomyocardial biopsy (EMB). EMB has potential complications and noted limitations, including interobserver variability in interpretation. Additional tests, such as basic cardiac biomarkers, cardiac imaging, gene expression profiling (GEP) scores, donor-derived cell-free DNA (dd-cfDNA) and the novel molecular microscope diagnostic system (MMDx) have become critical tools in rejection surveillance beyond standard EMB. METHODS: This paper describes an illustrative case followed by a review of MMDx within the context of other noninvasive screening modalities for rejection. CONCLUSIONS: We suggest MMDx be used to assist with early detection of rejection in cases of discordance between EMB and other noninvasive studies.

Stroke and Mechanical Circulatory Support in Adults.

PURPOSE OF THE REVIEW: Short-term and durable mechanical circulatory support (MCS) devices represent life-saving interventions for patients with cardiogenic shock and end-stage heart failure. This review will cover the epidemiology, risk factors, and treatment of stroke in this patient population. RECENT FINDINGS: Short-term devices such as intra-aortic balloon pump, Impella, TandemHeart, and Venoatrial Extracorporal Membrane Oxygenation, as well as durable continuous-flow left ventricular assist devices (LVADs), improve cardiac output and blood flow to the vital organs. However, MCS use is associated with high rates of complications, including ischemic and hemorrhagic strokes which carry a high risk for death and disability. Improvements in MCS technology have reduced but not eliminated the risk of stroke. Mitigation strategies focus on careful management of anti-thrombotic therapies. While data on therapeutic options for stroke are limited, several case series reported favorable outcomes with thrombectomy for ischemic stroke patients with large vessel occlusions, as well as with reversal of anticoagulation for those with hemorrhagic stroke. Stroke in patients treated with MCS is associated with high morbidity and mortality. Preventive strategies are targeted based on the specific form of MCS. Improvements in the design of the newest generation device have reduced the risk of ischemic stroke, though hemorrhagic stroke remains a serious complication.

Validity of echocardiography for detection of left ventricular thrombus with surgical validation in patients awaiting durable left ventricular assist device.

OBJECTIVE: Unrecognized left ventricular thrombi (LVT) can have devastating clinical implications and precludes patients with end-stage heart failure from undergoing left ventricular assist device (LVAD) implantation without cardiopulmonary bypass assistance. We assessed the reliability of an echocardiogram to diagnose LVT in patients with end-stage heart disease who underwent LVAD implantation. METHODS: A single-center retrospective study evaluated 232 consecutive adult patients requiring implantation of durable LVADs between 2005 and 2019. The validity of preoperative transthoracic echocardiogram (TTE) and intraoperative transesophageal echocardiogram (TEE) for diagnosing LVT was compared to direct inspection at the time of LVAD implantation. RESULTS: There were 232 patients that underwent LVAD implantation, with 226 patients (97%) receiving a preoperative TTE. Of those 226 patients, 32 patients (14%) received ultrasound enhancing agents (UEA). Intraoperative TEE images were available in 195 patients (84%). The sensitivity of TTE without UEA was 22% and specificity was 90% for detecting LVT, compared to 50% and 86%, respectively, for TTE with UEA. For intraoperative TEE, the sensitivity and specificity were 46% and 96%, respectively. The false omission rate ranged from 4% to 8% for all modalities of echocardiography. CONCLUSION: Among patients undergoing LVAD implantation, preoperative TTE and intraoperative TEE had poor sensitivity for LVT detection. Up to 8% of echocardiograms were incorrectly concluded to be negative for LVT on surgical validation. The low sensitivity and positive predictive value for diagnosing LVT suggest that echocardiography has limited reliability in this cohort of patients who are at high risk of LVT formation and its subsequent complications.

Clinical Utility of the Molecular Microscope Diagnostic System in a Real-World Transplant Cohort: Moving Towards a New Paradigm.

Objectives: To evaluate the clinical implications of adjunctive molecular gene expression analysis (MMDx ) of biopsy specimens in heart transplant (HT ) recipients with suspected rejection. Introduction: Histopathological evaluation remains the standard method for rejection diagnosis in HT. However, the wide interobserver variability combined with a relatively common incidence of "biopsy-negative" rejection has raised concerns about the likelihood of false-negative results. MMDx, which uses gene expression to detect early signs of rejection, is a promising test to further refine the assessment of HT rejection. Methods: Single-center prospective study of 418 consecutive for-cause endomyocardial biopsies performed between November 2022 and May 2024. Each biopsy was graded based on histology and assessed for rejection patterns using MMDx. MMDx results were deemed positive if borderline or definitive rejection was present. The impact of MMDx results on clinical management was evaluated. Primary outcomes were 1-year survival and graft dysfunction following MMDx-guided clinical management. Secondary outcomes included changes in donor-specific antibodies, MMDx gene transcripts, and donor-derived cell-free DNA (dd-cfDNA) levels. Results: We analyzed 418 molecular samples from 237 unique patients. Histology identified rejection in 32 cases (7.7%), while MMDx identified rejection in 95 cases (22.7%). Notably, in 79 of the 95 cases where MMDx identified rejection, histology results were negative, with the majority of these cases being antibody-mediated rejection (62.1%). Samples with rejection on MMDx were more likely to show a combined elevation of dd-cfDNA and peripheral blood gene expression profiling than those with borderline or negative MMDx results (36.7% vs 28.0% vs 10.3%; p<0.001). MMDx results led to the implementation of specific antirejection protocols or changes in immunosuppression in 20.4% of cases, and in 73.4% of cases where histology was negative and MMDx showed rejection. 1-year survival was better in the positive MMDx group where clinical management was guided by MMDx results (87.0% vs 78.6%; log rank p=0.0017). Conclusions: In our cohort, MMDx results more frequently indicated rejection than histology, often leading to the initiation of antirejection treatment. Intervention guided by positive MMDx results was associated with improved outcomes.

Relapsing Polychondritis Requiring Orthotopic Heart Transplant Despite Coronary Artery Bypass and Surgical Aortic Valve Replacement.

A 32-year-old man with a history of relapsing polychondritis presented with acute coronary syndrome due to aortitis with ostial coronary artery involvement from his underlying autoimmune condition. Concomitant aortic insufficiency with ostial coronary lesions is a rare complication of relapsing polychondritis, requiring a multidisciplinary team approach for management. (Level of Difficulty: Advanced.).

Angiotensin Converting Enzyme Inhibitor and Angiotensin II Receptor Blocker Use Among Outpatients Diagnosed With COVID-19.

Coronavirus disease 2019 (COVID-19) is a viral pandemic precipitated by the severe acute respiratory syndrome coronavirus 2. Since previous reports suggested that viral entry into cells may involve angiotensin converting enzyme 2, there has been growing concern that angiotensin converting enzyme inhibitor (ACEI) and angiotensin II receptor blocker (ARB) use may exacerbate the disease severity. In this retrospective, single-center US study of adult patients diagnosed with COVID-19, we evaluated the association of ACEI/ARB use with hospital admission. Secondary outcomes included: ICU admission, mechanical ventilation, length of hospital stay, use of inotropes, and all-cause mortality. Propensity score matching was performed to account for potential confounders. Among 590 unmatched patients diagnosed with COVID-19, 78 patients were receiving ACEI/ARB (median age 63 years and 59.7% male) and 512 patients were non-users (median age 42 years and 47.1% male). In the propensity matched population, multivariate logistic regression analysis adjusting for age, gender and comorbidities demonstrated that ACEI/ARB use was not associated with hospital admission (OR 1.2, 95%CI 0.5 to 2.7, p = 0.652). CAD and CKD/end stage renal disease [ESRD] remained independently associated with admission to hospital. All-cause mortality, ICU stay, need for ventilation, and inotrope use was not significantly different between the 2 study groups. In conclusion, among patients who were diagnosed with COVID-19, ACEI/ARB use was not associated with increased risk of hospital admission.

Dual Chamber Pacemaker Implantation Complicated by Left Anterior Descending Coronary Artery Injury.

A 53-year-old female underwent dual-chamber pacemaker implantation for tachy-brady syndrome, which was complicated by anterior ST-segment elevation myocardial infarction and ventricular fibrillation due to right ventricular lead impingement on the left anterior descending coronary artery. Coronary artery injury is a rare complication of cardiac device implantation which requires a multidisciplinary team for management. (Level of Difficulty: Beginner.).

Behavioral and brain temperature responses to salient environmental stimuli and intravenous cocaine in rats: effects of diazepam.

RATIONALE: While diazepam is an effective anxiolytic and somnolent drug in humans, its physiological and behavioral effects in animals are often variable. Differences in basal activity state (basal arousal) may be important in determining both this response variability and the pattern of drug influence on behavioral and physiological responses to natural arousing stimuli and other drugs. OBJECTIVES: To evaluate the changes in brain, muscle, and skin temperatures, and in locomotion induced in rats by several arousing stimuli and intravenous (i.v.) cocaine; and to assess how these responses are modulated by diazepam at a relatively low dose (1 mg/kg, i.p.). MATERIALS AND METHODS: Male rats were implanted with thermal probes in the nucleus accumbens (NAcc), temporal muscle, and subcutaneously, and equipped with a chronic i.v. catheter. They were exposed to 1-min tail-pinch, 1-min social interaction with another male and cocaine (1 mg/kg, i.v.) after administration of diazepam or saline. RESULTS: While the injection of either diazepam or saline resulted in similar locomotor activation and temperature responses, diazepam decreased basal brain and muscle temperatures for about 3 h; the temperature-decreasing effect of diazepam was oppositely related to basal brain temperature (r = -0.51). After diazepam, rats also showed weaker temperature and locomotor responses to both arousing stimuli; the effect was stronger for tail-pinch and for absolute temperature increases than relative changes. Although diazepam significantly decreased cocaine-induced locomotor activation, it had virtually no effects on cocaine-induced temperature responses in all locations. CONCLUSIONS: In accordance with the "law of initial values", the temperature-increasing effects of all tested arousing stimuli and temperature-decreasing effect of diazepam depend upon basal brain temperature. The greatest temperature effects are seen with arousing stimuli at low basal arousal (increases) and with diazepam at high basal arousal (decreases). This is a likely explanation for the variability seen with the physiological and behavioral effects of diazepam in animals.

Procedure of rectal temperature measurement affects brain, muscle, skin, and body temperatures and modulates the effects of intravenous cocaine.

Rectal probe thermometry is commonly used to measure body core temperature in rodents because of its ease of use. Although previous studies suggest that rectal measurement is stressful and results in long-lasting elevations in body temperatures, we evaluated how this procedure affects brain, muscle, skin, and core temperatures measured with chronically implanted thermocouple electrodes in rats. Our data suggest that the procedure of rectal measurement results in powerful locomotor activation, rapid and strong increases in brain, muscle, and deep body temperatures, as well as a biphasic, down-up fluctuation in skin temperature, matching the response pattern observed during tail-pinch, a representative stressful procedure. This response, moreover, did not habituate after repeated day-to-day testing. Repeated rectal probe insertions also modified temperature responses induced by intravenous cocaine. Under quiet resting conditions, cocaine moderately increased brain, muscle, and deep body temperatures. However, during repeated rectal measurements, which increased temperatures, cocaine induced both hyperthermic and hypothermic responses. Direct comparisons revealed that body temperatures measured by a rectal probe are typically lower (approximately 0.6 degrees C) and more variable than body temperatures recorded by chronically implanted electrodes; the difference is smaller at low and greater at high basal temperatures. Because of this difference and temperature increases induced by the rectal probe per se, cocaine had no significant effect on rectal temperatures compared to control animals exposed to repeated rectal probes. Therefore, although rectal temperature measurements provide a decent correlation with directly measured deep body temperatures, the arousing influence of this procedure may drastically modulate the effects of other arousing stimuli and drugs.

Impact of Methodologic Differences in Three-Dimensional Echocardiographic Measurements of the Aortic Annulus Compared with Computed Tomographic Angiography Before Transcatheter Aortic Valve Replacement.

BACKGROUND: Three-dimensional (3D) echocardiographic (3DE) imaging is an alternative to multi-detector row computed tomography (MDCT) for aortic annular measurement before transcatheter aortic valve replacement (TAVR). A commonly used direct planimetry from a reconstructed short-axis view has not been compared with semiautomated 3DE methods. Typically accepted optimal cutoffs for percent prosthesis-area oversizing of the balloon-expandable SAPIEN or SAPIEN XT valve to native annular size are approximately 5% to 15%. The aim of this study was to compare semiautomated and direct planimetric 3DE methods for aortic annular sizing with a gold standard of MDCT to determine predictive value for paravalvular regurgitation (PVR) and balloon postdilatation. METHODS: In this retrospective analysis, aortic annular cross-sectional area was measured from pre-TAVR imaging using (1) MDCT (CT_Area), (2) a 3D transesophageal echocardiographic (TEE) semiautomated method (3DE_Area_SA), and (3) a 3D TEE direct planimetric method (3DE_Area_Direct). Annular area percent oversizing was calculated. PVR after TAVR was assessed from intraoperative TEE imaging. Need for balloon postdilatation was recorded. RESULTS: One hundred patients who underwent TAVR with either the SAPIEN or SAPIEN XT balloon-expandable prosthesis were analyzed. Twenty-three patients had mild or greater PVR after TAVR. CT_Area was 442 ± 79 mm2, 3DE_Area_SA was 435 ± 81 mm2, and 3DE_Area_Direct was 429 ± 82 mm2. Both 3DE_Area_SA and 3DE_Area_Direct underestimated MDCT (P < .05). All methods were highly correlative (R = 0.88-0.93, P < .0001). Percent oversizing obtained by the three methods significantly predicted mild or greater PVR and need for balloon postdilatation by receiver operating characteristic analysis, with optimal cutoffs for CT_Area (9%-10%) and 3DE_Area_SA (14%) within the recommended ranges for the studied transcatheter valves and for 3DE_Area_Direct higher than the recommended range (18%-19%). Inter- and intraobserver reproducibility were lowest for 3DE_Area_Direct. CONCLUSIONS: Caution must be used when using 3D TEE direct planimetry of the aortic annulus, as optimal percent oversizing ranges approach the level associated with root injury, and measurements are less reproducible. Therefore, semiautomated 3DE planimetry is preferred to 3DE direct planimetry for aortic annulus sizing.

Practical determination of aortic valve calcium volume score on contrast-enhanced computed tomography prior to transcatheter aortic valve replacement and impact on paravalvular regurgitation: Elucidating optimal threshold cutoffs.

BACKGROUND: The threshold for the optimal computed tomography (CT) number in Hounsfield Units (HU) to quantify aortic valvular calcium on contrast-enhanced scans has not been standardized. Our aim was to find the most accurate threshold to predict paravalvular regurgitation (PVR) after transcatheter aortic valve replacement (TAVR). METHODS: 104 patients who underwent TAVR with the CoreValve prosthesis were studied retrospectively. Luminal attenuation (LA) in HU was measured at the level of the aortic annulus. Calcium volume score for the aortic valvular complex was measured using 6 threshold cutoffs (650 HU, 850 HU, LA × 1.25, LA × 1.5, LA+50, LA+100). Receiver-operating characteristic (ROC) analysis was performed to assess the predictive value for > mild PVR (n = 16). Multivariable analysis was performed to determine the accuracy to predict > mild PVR after adjustment for depth and perimeter oversizing. RESULTS: ROC analysis showed lower area under the curve (AUC) values for fixed threshold cutoffs (650 or 850 HU) compared to thresholds relative to LA. The LA+100 threshold had the highest AUC (0.81), and AUC was higher than all studied protocols, other than the LA x 1.25 and LA + 50 protocols, where the difference approached statistical significance (p = 0.05, and 0.068, respectively). Multivariable analysis showed calcium volume determined by the LAx1.25, LAx1.5, LA+50, and LA+ 100 HU protocols to independently predict PVR. CONCLUSIONS: Calcium volume scoring thresholds which are relative to LA are more predictive of PVR post-TAVR than those which use fixed cutoffs. A threshold of LA+100 HU had the highest predictive value.

The diagnosis and treatment of osteoporosis in men on androgen deprivation therapy for advanced carcinoma of the prostate.

PURPOSE: Androgen deprivation therapy (ADT) for advanced prostate cancer increases the risk of osteoporosis. Thus, the practicing urologist should be aware of the appropriate assessment and management. In this article we review the tests designed to diagnose osteoporosis as well as treatment options. We also review methods to monitor the response to therapy and make recommendations for management. MATERIALS AND METHODS: We performed a MEDLINE (1966 to 2004) search for the terms male, osteoporosis, bone mineral density, prostate cancer, androgen deprivation therapy, bisphosphonates, estrogen and the combinations thereof. We then constructed a management algorithm based on the best evidence available. RESULTS: Dual energy x-ray absorptiometry of the hip is the gold standard test for osteoporosis. Biochemical markers of bone turnover are not suitable for diagnosis but they have been shown to be useful for monitoring the response to treatment. Smoking cessation, weight bearing exercise, and vitamin D and calcium have been shown to help improve bone mineral density (BMD). Bisphosphonates have been demonstrated to increase BMD and decrease fracture risk in men with osteoporosis. Estrogens have also recently been shown to decrease bone turnover and increase BMD in men on ADT. CONCLUSIONS: Hip dual energy x-ray absorptiometry should be performed in all men who are anticipated to be on long-term ADT. In addition, all men on ADT should receive vitamin D and calcium supplementation, and perform regular weight bearing exercise. The value of smoking cessation cannot be overstated. In men who have osteopenia or osteoporosis bisphosphonate therapy should be initiated. Estrogen therapy has shown promise but specific recommendations cannot be made at this time.

CD40-mediated p38 mitogen-activated protein kinase activation is required for immunoglobulin class switch recombination to IgE.

BACKGROUND: Signaling through CD40 activates multiple kinases and signal pathways that drive diverse CD40-mediated biologic functions. The specific pathways activated by CD40 signaling involving CD40-dependent Ig class switch recombination (CSR) have not been defined. OBJECTIVE: We sought to dissect CD40-activated signaling required for CD40-mediated Ig CSR by using the specific signal pathway inhibitors, with the emphasis on CD40-activated p38 mitogen-activated protein kinase (p38 MAPK) signaling in CD40-mediated CSR to IgE. METHODS: Human B cells were costimulated with IL-4 plus anti-CD40 in the presence or absence of specific signal pathway inhibitors. Ig production, kinase phosphorylation, IgH epsilon germline transcripts and Smu-Sepsilon recombination were examined, and their relationships were analyzed. RESULTS: CD40-dependent IgE induction was inhibited by the specific p38 MAPK inhibitor SB203580 but not by the extracellular signal-regulated protein kinase-specific inhibitor PD98059 or the phosphatidylinositol 3-kinase-specific inhibitor LY294002. CD40 activation of p38 MAPK correlated with CD40-dependent IgE production, and IgE suppression by SB203580 correlated with the inhibition of CD40-activated p38 MAPK phosphorylation. Suppression of IgE production by SB203580 was not due to inhibition of cell proliferation because SB203580 did not suppress IL-4 plus alpha-CD40-induced cell proliferation. SB203580, but neither PD98059 nor LY294002, inhibited CD40-dependent Smu-Sepsilon recombination, as determined by using a digestion circularization PCR assay. The inhibitory effects of SB203580 on IgE production and Smu-Sepsilon recombination were directly related to its ability to suppress production of Ig epsilon germline transcripts. CONCLUSION: These results demonstrate that p38 MAPK is required for CD40-mediated class switching to IgE.

Protein kinase D is a downstream target of protein kinase Ctheta.

Protein kinase D (PKD/PKCmu immunoprecipitated from either COS-7 cells or Jurkat T lymphocytes transiently transfected with a constitutively active mutant of PKCtheta AE (PKCthetaAE) exhibited a marked increase in basal activity. In contrast, coexpression of constitutively active mutant of PKCzeta does not induce PKD activation in both types of cells. PKCthetaAE does not induce kinase activity in immunocomplexes of PKD kinase-deficient mutants PKDK618N or PKDD733A. PKD activation in response to PKCthetaAE signaling was completely prevented by treatment with the protein kinase C (PKC) inhibitors, GF I or Ro 31-8220, or by mutation of Ser-744 and Ser-748 to Ala in the kinase activation loop of PKD. Our results show that PKD is a downstream target of the theta isoform of PKC in both COS-7 cells and lymphocytes. The regulation of PKD by PKCtheta reveals a new pathway in the signaling network existing between multiple members of the PKC superfamily and PKD.

Proteome analysis of lipid rafts in Jurkat cells characterizes a raft subset that is involved in NF-kappaB activation.

Lipid rafts are detergent-insoluble membrane domains that play a key role in signal transduction by the T-cell antigen receptor. Proteome analysis revealed the presence of amidosulfobetaine-soluble signal transducing, integral membrane, cytoskeletal, heat shock, and GTP-binding proteins in rafts prepared from Jurkat cells. Several of these proteins were recruited to rafts by CD3/CD28 costimulation. Of particular interest is the inducible association of activated IkappaB kinase complexes with raft vesicles that could be captured with anti-flotillin-1 antibodies. Following amidosulfobetaine solubilization, flotillin-beta and IKKbeta underwent reciprocal co-immunoprecipitation. Treatment of Jurkat cells with methyl-beta-cyclodextrin disrupted the assembly and activation of this raft complex and also interfered in CD3/ CD28-induced activation of a NF-kappaB response element in the IL-2 promoter.