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Tunneling nanotubes (TNTs) are F‐actin‐based open‐ended tubular extensions that form following stresses, such as nutritional deprivation and oxidative stress. The chemotherapy agent 5‐fluorouracil (5‐FU) represents a significant stressor to cancer cells and induces thymidine deficiency, a state similar to nutritional deprivation. However, the ability of 5‐FU to induce TNT formation in cancer cells and potentially enhance survival has not been explored. In this study, we examined whether 5‐FU can induce TNT formation in MCF‐7 breast cancer cells. Cytotoxic doses of 5‐FU (150–350 μm) were observed to significantly induce TNT formation beginning at 24 h after exposure. TNTs formed following 5‐FU treatment probably originated as extensions of gap junctions as MCF‐7 cells detach from cell clusters. TNTs act as conduits for exchange of cellular components and we observed mitochondrial exchange through TNTs following 5‐FU treatment. 5‐FU‐induced TNT formation was inhibited by over 80% following treatment with the F‐actin‐depolymerizing agent, cytochalasin B (cytoB). The inhibition of TNTs by cytoB corresponded with increased 5‐FU‐induced cytotoxicity by 30–62% starting at 48 h, suggesting TNT formation aides in MCF‐7 cell survival against 5‐FU. Two other widely used chemotherapy agents, docetaxel and doxorubicin induced TNT formation at much lower levels than 5‐FU. Our work suggests that the therapeutic targeting of TNTs may increase 5‐FU chemotherapy efficacy and decrease drug resistance in cancer cells, and these findings merits further investigation.

Mitochondria oscillate along a morphological continuum from fragmented individual units to hyperfused tubular networks. Their position at the junction of catabolic and anabolic metabolism couples this morphological plasticity, called mitochondrial dynamics, to larger cellular metabolic programs, which in turn implicate mitochondria in a number of disease states. In many cancers, fragmented mitochondria engage the cell with the biosynthetic capacity of aerobic glycolysis in service of proliferation and progression. Chemo-resistant cancers, however, favor remodeling dynamics that yield fused mitochondrial assemblies utilizing oxidative phosphorylation (OXPHOS) through the electron transport chain (ETC). In this study, expression of Mitofusin-2 (MFN-2), a GTPase protein mediator of mitochondrial fusion, was found to closely correlate to Jurkat leukemia cell survival post doxorubicin (DxR) assault. Moreover, this was accompanied by dramatically increased expression of OXPHOS respiratory complexes and ATP Synthase, as well as a commensurate escalation of state III respiration and respiratory control ratio (RCR). Importantly, CRISPR knockout of MFN-2 resulted in a considerable decrease of doxorubicin (DxR) median lethal dose compared to a treated wildtype control, suggesting an important role of mitochondrial fusion in chemotherapy sensitivity and acute resistance.

Zavattaro, S. M., & Orr, S. K. (2017). Reflections on academic lives: Identifies, struggles, and triumphs in graduate school and beyond. New York, NY: Palgrave Macmillan. $139.99 (hardback), ISBN: 978-1-137-60008-0

Statistical learning (SL) is the ability to rapidly track statistical regularities and learn patterns in the environment. Recent studies show that SL is constrained by domain-specific features, rather than being a uniform learning mechanism across domains and modalities. This domain-specificity has been reflected at the neural level, as SL occurs in regions primarily involved in processing of specific modalities or domains of input. However, our understanding of how SL is constrained by domain-specific features in the developing brain is severely lacking. The present study aims to identify the functional neural profiles of auditory SL of linguistic and nonlinguistic regularities among children. Thirty children between 5 and 7 years old completed an auditory fMRI SL task containing interwoven sequences of structured and random syllable/tone sequences. Using traditional group univariate analyses and a group-constrained subject-specific analysis, frontal and temporal cortices showed significant activation when processing structured versus random sequences across both linguistic and nonlinguistic domains. However, conjunction analyses failed to identify overlapping neural indices across domains. These findings are the first to compare brain regions supporting SL of linguistic and nonlinguistic regularities in the developing brain and indicate that auditory SL among developing children may be constrained by domain-specific features., Competing Interests: Competing Interests: The authors have declared that no competing interests exist., (© 2024 Massachusetts Institute of Technology.)

Purpose: Exposomes are critical drivers of carcinogenesis. However, how they modulate tumor behavior remains unclear. Extensive clinical data link cigarette smoke as a key exposome that promotes aggressive tumors, higher rates of metastasis, reduced response to chemoradiotherapy, and suppressed anti-tumor immunity. We sought to determine whether smoke itself can modulate aggressive tumor behavior in head and neck squamous cell carcinoma (HNSCC) through reprogramming the cellular reductive state., Experimental Design: Using established human and murine HNSCC cell lines and syngeneic mouse models, we utilized conventional western blotting, steady state and flux metabolomics, RNA sequencing, quantitative proteomics and flow cytometry to analyze the impact of smoke exposure on HNSCC tumor biology., Results: Cigarette smoke persistently activated Nrf2 target genes essential for maintenance of the cellular reductive state and survival under conditions of increased oxidative stress in HNSCC regardless of HPV status. In contrast to e-cigarette vapor, conventional cigarette smoke mobilizes cellular metabolism toward oxidative stress adaptation, resulting in development of cross-resistance to cisplatin. In parallel, smoke exposure modulates both expression of PDL1 and the secretory phenotype of HNSCC cells through activation of NF-κB resulting in an altered tumor immune microenvironment (TIME) in syngeneic mouse models and altered PBMC differentiation that includes downregulated expression of antigen presentation and costimulatory genes in myeloid cells., Conclusion: Cigarette smoke exposome is a potent activator of the Nrf2 pathway and is a likely primary trigger for the tripartite phenotype of aggressive HNSCC consisting of: 1) reduced chemotherapy sensitivity, 2) enhanced metastatic potential and 3) suppressed anti-tumor immunity., Statement of Significance: The smoke exposome drives aggressive tumor behavior, treatment resistance and suppressed immunity through coordinated metabolic reprogramming. Successfully targeting this adaptation is critical to improving survival in smokers with head and neck cancer.

Background: The successful application of Machine Learning (ML) to many clinical problems can lead to its implementation as a medical device (MD), which is important to assess the associated risks., Methods: An anemia control model (ACM), certified as MD, may face adverse events as a result of wrong predictions that are translated into suggestions of doses of erythropoietic stimulating agents to dialysis patients. Risks are assessed as the combination of severity and probability of a given hazard. While severities are typically assessed by clinicians, probabilities are tightly related to the performance of the predictive model., Results: A postmarketing data set formed by all adult patients registered in French, Portuguese, and Spanish clinics, belonging to an international network, was considered; 3876 patients and 11,508 suggestions were eventually included. The achieved results show that there are no statistical differences between the probabilities of adverse events that are estimated in the ACM test set (using only Spanish clinics) and those actually observed in the postmarketing cohort., Conclusions: The risks of an ACM-MD can be accurately and robustly estimated, thus enhancing patients' safety. The proposed methodology is applicable to other clinical decisions based on predictive models since our proposal does not depend on the particular predictive model.

Objective: This study aims to reconstruct a real-world Crash Injury Research and Engineering Network vehicle-to-pedestrian impact to supplement the determination of pedestrian kinematics and injury causation., Methods: A case involving a 46-year-old male pedestrian with a height of 163 cm and mass of 100 kg that was impacted by a 2019 Dodge Charger Pursuit police cruiser at an approximate velocity of 20.1 m/s was reconstructed. The case vehicle was represented by a rigid shell of a 2019 Dodge Charger vehicle exterior from an open-source database. The case pedestrian was represented by the Global Human Body Models Consortium (GHBMC) 50th percentile male simplified pedestrian human body model. The GHBMC model was isometrically scaled to a height of 163 cm and the external layer of flesh was morphed to a male reference geometry with the same age, height, and body mass index as the case pedestrian. The approximate location and position of the pedestrian at the time of impact was determined from case vehicle dashboard camera images and the pedestrian model was adjusted accordingly., Results: Reconstruction kinematics aligned with proposed CIREN case kinematics. The GHBMC model predicted fractures of the left inferior ischiopubic ramus, superior pubic ramus, ilium, sacral ala, acetabulum, and right ilium., Conclusions: Finite element reconstructions of real-world pedestrian impacts are useful for analyzing pedestrian kinematics and provide an effective tool for improving pedestrian impact injury analyses.

Objective: Finite element human body models (HBMs) must be certified for use within the EuroNCAP pedestrian safety assessment protocol. We demonstrate that the Global Human Body Model Consortium (GHBMC) simplified pedestrian series of HBMs meet all criteria set forth in Technical Bulletin (TB) 024 (v 1.1 Jan. 2019) for model certification. We further explore variation in head contact time (HIT) and location by HBM size and impact speed across 48 full body impact simulations. Methods: The EuroNCAP Pedestrian Protocol (v. 8.5, Oct. 2018) assesses the overall safety of adult and child pedestrians by outlining a variety of physical tests and finite element simulations using HBMs. These tests are designed to assess the efficacy of vehicle safety technology such as active bonnets. The 50th percentile male simplified pedestrian model (M50-PS, H:175 cm, W:74.5 kg), six-year-old (6YO-PS, H:117 cm, W:23.4 kg), 5th percentile female (F05-PS, H:150 cm, W:50.7 kg), and 95th percentile male (M95-PS, H:190 cm, W:102 kg) were simulated through the suite of cases totaling 48 simulations (12 each). The process gathers three kinematic trajectories and contact force data from designated anatomical locations. The impacting vehicles include a family car (FCR), multi-purpose vehicle (MPV), roadster (RDS), and sports utility vehicle (SUV), all provided by TU Graz, Vehicle Safety Institute as part of the Coherent Project, each simulated at 30 kph, 40 kph, and 50 kph. Each simulation underwent a 23-point pre-simulation check and post-simulation model response comparison. The posture of all HBMs met criteria consisting of 15 measures. All simulations were conducted in LS-Dyna R. 7.1.2. Results and Conclusions: All simulations normal terminated. For each of the simulations, sagittal plane coordinate histories of the center of the head, 12th thoracic vertebrae, and center of acetabulum were compared with standard corridors and did not exceed the tolerance of 50 mm deviation. Head contact time was also compared with the reference values and did not exceed the tolerance interval of +3.5% and -7%. Comparison of contact forces was required for monitoring purposes only. The head contact time of the models for each simulation was recorded and compared by model size, impact speed, and vehicle geometry. Head contact times varied by roughly 3-fold, were lowest for the child model, and showed the greatest sensitivity for the tallest stature model (M95-PS). As stated in the certification process, other body sizes within a model family qualify for certification if the 50th percentile male model passes, provided that model sizes meet the required posture.

Interleukin-7 is a cytokine essential for T cell homeostasis. IL-7 binds to cellular IL-7 receptors in competition with a soluble form of the receptor (sIL-7Rα). We hypothesized that altered sIL-7Rα levels may cause adverse outcomes in patients undergoing HSCT. In parallel, we investigated the impact of the IL-7Rα SNP rs6897932, which has been associated with release of IL-7R. The sIL-7Rα levels decreased during HSCT (from 114ng/ml before to 48ng/ml at day +14 (P<0.0001)). This pattern was inversely mirrored by IL-7. The IL-7/sIL-7Rα ratio at day +14 was significantly higher in patients developing grades II-IV aGVHD (OR=4.3, P=0.026). Furthermore, donor carriage of the rs6897932 T allele was associated with reduced sIL-7Rα levels, increased risk of grades II-IV aGVHD (OR=2.4, P=0.055) and increased transplant-related mortality (CC=4.5%, CT=21.4% and TT=27.3%, P=0.0037). In conclusion, this study suggests an impact of sIL-7Rα levels and rs6897932 donor genotype on alloreactivity and outcome after HSCT., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Objective: The significant computational resources required to execute detailed human body finite-element models has motivated the development of faster running, simplified models (e.g., GHBMC M50-OS). Previous studies have demonstrated the ability to modularly incorporate the validated GHBMC M50-O brain model into the simplified model (GHBMC M50-OS+B), which allows for localized analysis of the brain in a fraction of the computation time required for the detailed model. The objective of this study is to validate the head and neck kinematics of the GHBMC M50-O and M50-OS (detailed and simplified versions of the same model) against human volunteer test data in frontal and lateral loading. Furthermore, the effect of modular insertion of the detailed brain model into the M50-OS is quantified., Methods: Data from the Navy Biodynamics Laboratory (NBDL) human volunteer studies, including a 15g frontal, 8g frontal, and 7g lateral impact, were reconstructed and simulated using LS-DYNA. A five-point restraint system was used for all simulations, and initial positions of the models were matched with volunteer data using settling and positioning techniques. Both the frontal and lateral simulations were run with the M50-O, M50-OS, and M50-OS+B with active musculature for a total of nine runs., Results: Normalized run times for the various models used in this study were 8.4 min/ms for the M50-O, 0.26 min/ms for the M50-OS, and 0.97 min/ms for the M50-OS+B, a 32- and 9-fold reduction in run time, respectively. Corridors were reanalyzed for head and T1 kinematics from the NBDL studies. Qualitative evaluation of head rotational accelerations and linear resultant acceleration, as well as linear resultant T1 acceleration, showed reasonable results between all models and the experimental data. Objective evaluation of the results for head center of gravity (CG) accelerations was completed via ISO TS 18571, and indicated scores of 0.673 (M50-O), 0.638 (M50-OS), and 0.656 (M50-OS+B) for the 15g frontal impact. Scores at lower g levels yielded similar results, 0.667 (M50-O), 0.675 (M50-OS), and 0.710 (M50-OS+B) for the 8g frontal impact. The 7g lateral simulations also compared fairly with an average ISO score of 0.565 for the M50-O, 0.634 for the M50-OS, and 0.606 for the M50-OS+B. The three HBMs experienced similar head and neck motion in the frontal simulations, but the M50-O predicted significantly greater head rotation in the lateral simulation., Conclusion: The greatest departure from the detailed occupant models were noted in lateral flexion, potentially indicating the need for further study. Precise modeling of the belt system however was limited by available data. A sensitivity study of these parameters in the frontal condition showed that belt slack and muscle activation have a modest effect on the ISO score. The reduction in computation time of the M50-OS+B reduces the burden of high computational requirements when handling detailed HBMs. Future work will focus on harmonizing the lateral head response of the models and studying localized injury criteria within the brain from the M50-O and M50-OS+B.

An increasing number of studies suggest an important role of host immunity as a barrier to tumor formation and progression. Complex mechanisms and multiple pathways are involved in evading innate and adaptive immune responses, with a broad spectrum of chemicals displaying the potential to adversely influence immunosurveillance. The evaluation of the cumulative effects of low-dose exposures from the occupational and natural environment, especially if multiple chemicals target the same gene(s) or pathway(s), is a challenge. We reviewed common environmental chemicals and discussed their potential effects on immunosurveillance. Our overarching objective was to review related signaling pathways influencing immune surveillance such as the pathways involving PI3K/Akt, chemokines, TGF-β, FAK, IGF-1, HIF-1α, IL-6, IL-1α, CTLA-4 and PD-1/PDL-1 could individually or collectively impact immunosurveillance. A number of chemicals that are common in the anthropogenic environment such as fungicides (maneb, fluoxastrobin and pyroclostrobin), herbicides (atrazine), insecticides (pyridaben and azamethiphos), the components of personal care products (triclosan and bisphenol A) and diethylhexylphthalate with pathways critical to tumor immunosurveillance. At this time, these chemicals are not recognized as human carcinogens; however, it is known that they these chemicalscan simultaneously persist in the environment and appear to have some potential interfere with the host immune response, therefore potentially contributing to promotion interacting with of immune evasion mechanisms, and promoting subsequent tumor growth and progression., (© The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

The effect of superimposed ac and dc electric fields on the formation of electroconvection and flexoelectric patterns in nematic liquid crystals was studied. For selected ac frequencies, an extended standard model of the electrohydrodynamic instabilities was used to characterize the onset of pattern formation in the two-dimensional parameter space of the magnitudes of the ac and dc electric field components. Numerical as well as approximate analytical calculations demonstrate that depending on the type of patterns and on the ac frequency, the combined action of ac and dc fields may either enhance or suppress the formation of patterns. The theoretical predictions are qualitatively confirmed by experiments in most cases. Some discrepancies, however, seem to indicate the need to extend the theoretical description.

The naive phenotype of cord blood (CB) T cells may reduce graft-versus-host disease after umbilical cord blood transplantation, but this naivety and their low absolute numbers also delays immune reconstitution, producing higher infection-related mortality that is predominantly related to CMV, adenovirus (Adv), and EBV. Adoptive immunotherapy with peripheral blood-derived virus-specific cytotoxic T lymphocytes (CTLs) can effectively prevent viral disease after conventional stem cell transplantation, and we now describe the generation of single cultures of CTLs from CB that are specific for multiple viruses. Using EBV-infected B cells transduced with a clinical-grade Ad5f35CMVpp65 adenoviral vector as sources of EBV, Adv, and CMV antigens, we expanded virus-specific T cells even from CB T cells with a naive phenotype. After expansion, each CTL culture contained both CD8(+) and CD4(+) T-cell subsets, predominantly of effector memory phenotype. Each CTL culture also had HLA-restricted virus-specific cytotoxic effector function against EBV, CMV, and Adv targets. The CB CTLs recognized multiple viral epitopes, including CD4-restricted Adv-hexon epitopes and immunosubdominant CD4- and CD8-restricted CMVpp65 epitopes. Notwithstanding their naive phenotype, it is therefore possible to generate trivirus-specific CTLs in a single culture of CB, which may be of value to prevent or treat viral disease in CB transplant recipients. This study is registered at www.clinicaltrials.gov as NCT00078533.

Background: IL-7 is an essential cytokine in T-cell development and homeostasis. It binds to the IL-7R receptor, a complex of the IL-7Ralpha (CD127) and common gamma (CD132) chains. There is significant interest in evaluating the expression of CD127 on human T-cells as it often decreased in medical conditions leading to lymphopenia. Previous reports showed the usefulness of CD127 as a prognostic marker in viral infections such as HIV, CMV, EBV and HCV. A soluble CD127 (sCD127) is released in plasma and may contribute to disease pathogenesis through its control on IL-7 activities. Measuring sCD127 is important to define its role and may complement existing markers used in lymphopenic disease management. We describe a new quantitative assay for the measurement of sCD127 in plasma and report sCD127 concentrations in healthy adults., Methodology/principal Findings: We developed a quantitative bead-based sCD127 capture assay. Polyclonal CD127-specific antibodies were chosen for capture and a biotinylated monoclonal anti-CD127 antibody was selected for detection. The assay can detect native sCD127 and recombinant sCD127 which served as the calibrator. The analytical performance of the assay was characterized and the concentration and stability of plasma sCD127 in healthy adults was determined. The assay's range was 3.2-1000 ng/mL. The concentration of plasma sCD127 was 164+/-104 ng/mL with over a log variation between subjects. Individual sCD127 concentrations remained stable when measured serially during a period of up to one year., Conclusions/significance: This is the first report on the quantification of plasma sCD127 in a population of healthy adults. Soluble CD127 plasma concentrations remained stable over time in a given individual and sCD127 immunoreactivity was resistant to repeated freeze-thaw cycles. This quantitative sCD127 assay is a valuable tool for defining the potential role of sCD127 in lymphopenic diseases.

Background: Anaphylaxis is a serious allergic reaction that is rapid in onset and may cause death. Allergists and emergency department (ED) physicians recognize the need for multidisciplinary efforts to increase public awareness of anaphylaxis, improve patient education, and enhance emergency and long-term management for the millions of Americans at risk for anaphylactic reactions., Objectives: To provide an overview of the scientific literature documenting inconsistencies and limitations in the management of anaphylaxis in the ED, to highlight the need to enhance ED discharge instructions and patient education materials, and to introduce the SAFE system, an aid to improve the ongoing management and avoidance of factors that contribute to anaphylaxis., Methods: An expert panel of allergists and ED physicians was convened by the American College of Allergy, Asthma and Immunology and the American College of Emergency Physicians to develop recommendations for educational materials on anaphylaxis for ED personnel and patients., Results: The panel developed the acronym SAFE, a mnemonic device to remind physicians of the 4 basic action steps suggested for the care of a patient with anaphylaxis: (1) Seek support, (2) Allergen identification and avoidance, (3) Follow-up for specialty care, and (4) Epinephrine for emergencies., Conclusion: The SAFE system is designed as a tool to raise awareness of anaphylaxis and its treatments among individuals at risk and the ED personnel charged with their care. Future studies need to examine the impact of the SAFE system in decreasing the morbidity and mortality rates associated with anaphylaxis and other severe allergic reactions.

One factor limiting the therapeutic efficacy of cord blood (CB) hematopoietic progenitor cell (HPC) transplantation is the low cell dose of the graft. This is associated with an increased incidence of delayed or failed engraftment. Cell dose can be increased and the efficacy of CB transplantation potentially improved, by ex vivo CB expansion before transplantation. Two ex vivo CB expansion techniques were compared: (1) CD133+ selection followed by ex vivo liquid culture and (2) co-culture of unmanipulated CB with bone-marrow-derived mesenchymal stem cells (MSCs). Ex vivo culture was performed in medium supplemented with granulocyte colony-stimulating factor, stem cell factor and either thrombopoietin or megakaryocyte growth and differentiation factor. Expansion was followed by measuring total nucleated cell (TNC), CD133+ and CD34+ cell, colony-forming unit and cobblestone area-forming cell output. When compared to liquid culture, CB-MSC co-culture (i) required less cell manipulation resulting in less initial HPC loss and (ii) markedly improved TNC and HPC output. CB-MSC co-culture therefore holds promise for improving engraftment kinetics in CB transplant recipients.

Background: Umbilical cord blood (CB) is a promising source of hematopoietic stem cells for allogeneic transplantation. However, delayed engraftment and impaired immune reconstitution remain major limitations. Enrichment of donor grafts with CB T cells expanded ex vivo might facilitate improved T-cell immune reconstitution post-transplant. We hypothesized that CB T cells could be expanded using paramagnetic microbeads covalently linked to anti-CD3 and anti-CD28 Ab., Methods: CB units were divided into three fractions: (1) cells cultured without beads, (2) cells cultured with beads and (3) cells cultured with beads following CD3+ magnetic enrichment. All fractions were cultured for 14 days in the presence of IL-2 (200 IU/mL)., Results: A mean 100-fold expansion (range 49-154) of total nucleated cells was observed in the CD3+ magnetically enriched fraction. Following expansion, CB T cells retained a naive and/or central memory phenotype and contained a polyclonal TCR diversity demonstrated by spectratyping., Discussion: Our data provide evidence that naive and diverse CB T cells may be expanded ex vivo and warrant additional studies in the setting of human CB transplantation.

The monoclonal antibody (MAb) AR20.5 is a murine MAb, generated against the tandem repeat protein backbone of the tumor-associated antigen MUC1. MAb AR20.5 reacts strongly with either the soluble form or the cell surface epitope of MUC1 on many human cancer cell lines. It also reacts with a 23-amino acid MUC1 peptide, E23, which includes the core tandem repeat sequence. Epitope mapping confirmed that MAb AR20.5 recognizes a minimum of six residues with the sequence DTRPAP. Inhibition of glycosylation of MUC1 resulted in decreased binding of MAb AR20.5 to cell surface MUC1, suggesting that MAb AR20.5 binding is carbohydrate dependent. The antibody was studied in a human PBL-SCID/beige mouse model to evaluate its effect on progression of NIH:OVARCAR-3 tumors. Tumor reduction was observed in mice injected with MAb AR20.5, but not in mice treated with control murine antibody or PBS (p < 0.001 and p < 0.05, respectively). An anti-tumor effect could also be demonstrated in a CB6F1 mouse model with the MUC1 transfectoma 413BCR.

The channel-forming protein called VDAC forms the major pathway in the mitochondrial outer membrane and controls metabolite flux across that membrane. The different VDAC isoforms of a species may play different roles in the regulation of mitochondrial functions. The mouse has three VDAC isoforms (VDAC1, VDAC2 and VDAC3). These proteins and different versions of VDAC3 were expressed in yeast cells (S. cerevisiae) missing the major yeast VDAC gene and studied using different approaches. When reconstituted into liposomes, each isoform induced a permeability in the liposomes with a similar molecular weight cutoff (between 3,400 and 6,800 daltons based on permeability to polyethylene glycol). In contrast, electrophysiological studies on purified proteins showed very different channel properties. VDAC1 is the prototypic version whose properties are highly conserved among other species. VDAC2 also has normal gating activity but may exist in 2 forms, one with a lower conductance and selectivity. VDAC3 can also form channels in planar phospholipid membranes. It does not insert readily into membranes and generally does not gate well even at high membrane potentials (up to 80 mV). Isolated mitochondria exhibit large differences in their outer membrane permeability to NADH depending on which of the mouse VDAC proteins was expressed. These differences in permeability could not simply be attributed to different amounts of each protein present in the isolated mitochondria. The roles of these different VDAC proteins are discussed.

A series of compounds related to oxathiin carboxanilide has been identified as nonnucleoside reverse transcriptase inhibitors (NNRTIs) of HIV-1, and structure-activity relationships have been described (Buckheit RW, et al.: Antimicrob Agents Chemother 1995;39:2718-2727). Three new analogs (UC040, UC82, and UC781) inhibited laboratory and clinical isolates of HIV-1, including isolates representative of the various clades of HIV-1 found worldwide, in both established and fresh human cells. Virus isolates with the amino acid changes L100I, K103N, V106I, and Y181C in the reverse transcriptase were partially resistant to these compounds. However, UC781 inhibited these virus isolates at low nontoxic concentrations, presenting a broad in vitro therapeutic index. As with other NNRTIs, each of the compounds synergistically interacted with AZT to inhibit HIV-1 replication. UC781 possesses a favorable pharmacokinetic profile in mice with a high level of oral bioavailability. Plasma concentrations reached maximum levels within 2 to 4 hr of oral administration and remained in excess of those required for in vitro anti-HIV activity for at least 24 hr after a single oral dose. When evaluated in a murine hollow fiber implant model of HIV infection, UC781 dosed orally or parenterally was able to suppress HIV replication completely in this model system, providing evidence of the in vivo efficacy of the compound.

Albeit still unproven, it is supposed that the presence and number of asymptomatic circulating cerebral microemboli detected by transcranial Doppler ultrasound (TCD) may be an indicator of stroke risk. Little research has been done to assess the reproducibility of these data and the required time for recording. We examined one middle cerebral artery in 7 subjects with carotid, aortic or cardiac embolic source for 24 h by TCD. Analysis for embolic signals was done off line completely blinded to the diagnosis and the time of the day during the recording. Embolic signals were found in all 7 subjects varying from 0 per h to 13 per h. Embolic signals occurred throughout the day with a nonsignificant tendency towards higher values in the early morning hours. Half an hour recording would not have been suitable to rule out or to confirm the presence of embolic signals as there were too many gaps between embolic signals of more than 1/2 h. One hour is the required minimum. Concerning the number of embolic signals, even a recording of four hours yielded variabilities of 0.25 and 8 embolic signals per hour in the same person. We recommend to perform follow-up studies the same time of the day. In patients with a low number of embolic signals longer recordings or a lower detection threshold with a higher number of detected embolic signals are necessary to compare frequencies of embolic signals.

Previous studies demonstrated that human cell lines can be cultivated in hollow fibers in the subcutaneous and intraperitoneal compartments of mice. We have extended the range of cell lines to include cells infected with the human immunodeficiency virus (HIV). Furthermore, these HIV-infected cells have been shown to replicate in the hollow fibers located in both physiologic compartments (intraperitoneal and subcutaneous) of SCID mice. Treatment of the host mice with antiviral agents can suppress virus replication in these hollow fiber cultures. The potential use of this system for early in vivo screening of anti-HIV compounds is discussed.