Expression and function of ryanodine receptor related pathways in PCB tolerant Atlantic killifish (Fundulus heteroclitus) from New Bedford Harbor, MA, USA

Author Posting. © The Author(s), 2014. This is the author's version of the work. It is posted here by permission of Elsevier for personal use, not for redistribution. The definitive version was published in Aquatic Toxicology 159 (2015): 156-166, doi:10.1016/j.aquatox.2014.12.017.Atlantic killifish (Fundulus heteroclitus) thrive in New Bedford Harbor (NBH), MA, highly contaminated with polychlorinated biphenyls (PCBs). Resident killifish have evolved tolerance to dioxin-like (DL) PCBs, whose toxic effects through the aryl hydrocarbon receptor (AhR) are well studied. In NBH, non-dioxin like PCBs (NDL PCBs), which lack activity toward the AhR, vastly exceed levels of DL congeners yet how killifish counter NDL toxic effects has not been explored. In mammals and fish, NDL PCBs are potent activators of ryanodine receptors (RyR), Ca2+ release channels necessary for a vast array of physiological processes. In the current study we compared the expression and function of RyR related pathways in NBH killifish with killifish from the reference site at Scorton Creek (SC, MA). Relative to the SC fish, adults from NBH displayed increased levels of skeletal muscle RyR1 protein, and increased levels of FK506-binding protein 12 kDa (FKBP12), an accessory protein essential for NDL PCB-triggered changes in RyR channel function. In accordance with increased RyR1 levels, NBH killifish displayed increased maximal ligand binding, increased maximal response to Ca2+ activation and increased maximal response to activation by the NDL PCB congener PCB 95. Compared to SC, NBH embryos and larvae had increased levels of mtor and ryr2 transcripts at multiple stages of development, and generations, while levels of serca2 were decreased at 9 days post-fertilization in the F1 and F2 generations. These findings suggest that there are compensatory and heritable changes in RyR mediated Ca2+ signaling proteins or potential signaling partners in NBH killifish.Funding was provided through the NIEHS Superfund Research Program UC Davis (INP and EBF; P42-ES004699) and Boston University (JJS and JVG; P42-ES007381). Support was supplied via the UC Davis NHLBI Training Grant (T32-HL086350, EBF). Additional support came from NIEHS 1R01-ES014901, 1R01-ES017425, the UC Davis Center for Children’s Environmental Health (1P01-ES011269, U.S. Environmental Protection Agency Grant 8354320), and an unrestricted JB Johnson Foundation gift grant.2015-12-1

Modeling of bone conduction of sound in the human head using hp-finite elements: Code design and verification

We focus on the development of a reliable numerical model for investigating the bone-conduction of sound in the human head. The main challenge of the problem is the lack of fundamental knowledge regarding the transmission of acoustic energy through non-airborne pathways to the cochlea. A fully coupled model based on the acoustic/elastic interaction problem with a detailed resolution of the cochlea region and its interface with the skull and the air pathways, should provide an insight into this fundamental, long standing research problem. To this aim we have developed a 3D hp-finite element code that supports elements of all shapes (tetrahedra, prisms and pyramids) to better capture the geometrical features of the head. We have tested the code on a multilayered sphere and employed it to solve an idealized model of head. In the future we hope to attack a model with a more realistic geometry

Population genetic diversity and fitness in multiple environments

<p>Abstract</p> <p>Background</p> <p>When a large number of alleles are lost from a population, increases in individual homozygosity may reduce individual fitness through inbreeding depression. Modest losses of allelic diversity may also negatively impact long-term population viability by reducing the capacity of populations to adapt to altered environments. However, it is not clear how much genetic diversity within populations may be lost before populations are put at significant risk. Development of tools to evaluate this relationship would be a valuable contribution to conservation biology. To address these issues, we have created an experimental system that uses laboratory populations of an estuarine crustacean, <it>Americamysis bahia </it>with experimentally manipulated levels of genetic diversity. We created replicate cultures with five distinct levels of genetic diversity and monitored them for 16 weeks in both permissive (ambient seawater) and stressful conditions (diluted seawater). The relationship between molecular genetic diversity at presumptive neutral loci and population vulnerability was assessed by AFLP analysis.</p> <p>Results</p> <p>Populations with very low genetic diversity demonstrated reduced fitness relative to high diversity populations even under permissive conditions. Population performance decreased in the stressful environment for all levels of genetic diversity relative to performance in the permissive environment. Twenty percent of the lowest diversity populations went extinct before the end of the study in permissive conditions, whereas 73% of the low diversity lines went extinct in the stressful environment. All high genetic diversity populations persisted for the duration of the study, although population sizes and reproduction were reduced under stressful environmental conditions. Levels of fitness varied more among replicate low diversity populations than among replicate populations with high genetic diversity. There was a significant correlation between AFLP diversity and population fitness overall; however, AFLP markers performed poorly at detecting modest but consequential losses of genetic diversity. High diversity lines in the stressful environment showed some evidence of relative improvement as the experiment progressed while the low diversity lines did not.</p> <p>Conclusions</p> <p>The combined effects of reduced average fitness and increased variability contributed to increased extinction rates for very low diversity populations. More modest losses of genetic diversity resulted in measurable decreases in population fitness; AFLP markers did not always detect these losses. However when AFLP markers indicated lost genetic diversity, these losses were associated with reduced population fitness.</p

Regulation of pregnane-X-receptor, CYP3A and P-glycoprotein genes in the PCB-resistant killifish (Fundulus heteroclitus) population from New Bedford Harbor

Author Posting. © The Author(s), 2014. This is the author's version of the work. It is posted here by permission of Elsevier for personal use, not for redistribution. The definitive version was published in Aquatic Toxicology 159 (2015): 198-207, doi:10.1016/j.aquatox.2014.12.010.Killifish survive and reproduce in the New Bedford Harbor (NBH) in Massachusetts (MA), USA, a site severely contaminated with polychlorinated biphenyls (PCBs) for decades. Levels of 22 different PCB congeners were analyzed in liver from killifish collected in 2008. Concentrations of dioxin-like PCBs in liver of NBH killifish were ~400 times higher, and the levels of non-dioxin-like PCBs ~3000 times higher than in killifish from a reference site, Scorton Creek (SC), MA. The NBH killifish are known to be resistant to the toxicity of dioxin-like compounds and to have a reduced aryl hydrocarbon receptor (AhR) signaling response. Little is known about the responses of these fish to non-dioxin-like PCBs, which are at extraordinarily high levels in NBH fish. In mammals, some non-dioxin-like PCB congeners act through nuclear receptor 1I2, the pregnane-X-receptor (PXR). To explore this pathway in killifish, a PXR cDNA was sequenced and its molecular phylogenetic relationship to other vertebrate PXRs was determined. Killifish were also collected in 2009 from NBH and SC, and after four months in the laboratory they were injected with a single dose of either the dioxin-like PCB 126 (an AhR agonist) or the non-dioxin-like PCB 153 (a mammalian PXR agonist). Gills and liver were sampled three days after injection and transcript levels of PXR, cytochrome P450 3A (CYP3A), P-glycoprotein (Pgp), AhR2 and cytochrome P450 1A (CYP1A) were measured by quantitative PCR. As expected, there was little effect of PCB exposure on AhR2 or CYP1A in liver and gills of NBH fish. In NBH fish, but not in SC fish, there was increased expression of hepatic PXR, CYP3A and Pgp genes upon exposure to either of the two PCB congeners. However, basal PXR and Pgp mRNA levels in liver of NBH fish were significantly lower than in SC fish. A different pattern was seen in gills, where there were no differences in basal expression of these genes between the two populations. In SC fish, but not in NBH fish, there was increased expression of branchial PXR and CYP3A upon exposure to PCB126 and of CYP3A upon exposure to PCB153. The results suggest a difference between the two populations in non-AhR transcription factor signaling in liver and gills, and that this could involve killifish PXR. It also implies possible cross-regulatory interactions between that factor (presumably PXR) and AhR2 in liver of these fish.This study was supported by grants from FORMAS (216-2007-468) and University of Gothenburg to MCC, and by the Superfund Research Program at Boston University, NIH grant P42ES007381 to JJS, MEH, and SIK. Data interpretation was aided by reference to a preliminary draft of the Fundulus heteroclitus genome sequence, which was supported by funding from the National Science Foundation (collaborative research grants DEB-1120512, DEB-1265282, DEB-1120013, DEB-1120263, DEB-1120333, DEB-1120398). This study was also supported by NOAA Grant No. NA16RG2273 (WHOI Sea Grant Project No. R/P-70 to SIK and MEH) and by funding from Adlerbertska Forskningsstiftelsen, Helge Ax:son Johnsons Stiftelse and Wilhelm och Martina Lundgrens Vetenskapsfond to BW and JG

Proceedings from the Second Haploidentical Stem Cell Transplantation Symposium-Haplo2014, San Francisco, California, December 4, 2014

Significant progress has been made over the past decade in haploidentical transplantation, with the development of novel methods to control intense alloreactive reactions generated in the major HLA-mismatched setting. Application of post-transplantation cyclophosphamide has gained worldwide acceptance as an effective and low-cost way to perform this type of transplantation, with outcomes now similar to those from HLA-matched unrelated donors. These advances have resulted in improved treatment-related mortality, whereas disease relapse has emerged as the most common cause of treatment failure. In addition, improvements in immunologic reconstitution after transplantation are much needed, not only in haploidentical transplantation but in all forms of stem cell transplantation. This symposium has focused on some of the most promising methods to control alloreactivity in this form of transplantation and application of cellular therapy to prevent disease relapse after transplantation, as well as understanding immunologic reconstitution and foreseeable approaches to improve immune recovery after transplantation

Clofarabine ± Fludarabine with Once Daily i.v. Busulfan as Pretransplant Conditioning Therapy for Advanced Myeloid Leukemia and MDS

Although a combination of i.v. busulfan (Bu) and fludarabine (Flu) is a safe, reduced-toxicity conditioning program for acute myelogenous leukemia/myelodysplastic syndromes (AML/MDS), recurrent leukemia posttransplantation remains a problem. To enhance the conditioning regimen’s antileukemic effect, we decided to supplant Flu with clofarabine (Clo), and assayed the interactions of these nucleoside analogs alone and in combination with Bu in Bu-resistant human cell lines in vitro. We found pronounced synergy between each nucleoside and the alkylator but even more enhanced cytotoxic synergy when the nucleoside analogs were combined prior to exposing the cells to Bu. We then designed a 4-arm clinical trial in patients with myeloid leukemia undergoing allogeneic stem cell transplantation (allo-SCT). Patients were adaptively randomized as follows: Arm I–Clo:Flu 10:30 mg/m2, Arm II—20:20 mg/m2, Arm III—30:10 mg/m2, and Arm IV–single-agent Clo at 40 mg/m2. The nucleoside analog(s) were/was infused over 1 hour once daily for 4 days, followed on each day by Bu, infused over 3 hours to a pharmacokinetically targeted daily area under the curve (AUC) of 6000 μMol-min ± 10%. Fifty-one patients have been enrolled with a minimum follow-up exceeding 100 days. There were 32 males and 19 females, with a median age of 45 years (range: 6-59). Nine patients had chronic myeloid leukemia (CML) (BC: 2, second AP: 3, and tyrosine-kinase inhibitor refractory first chronic phase [CP]: 4). Forty-two patients had AML: 14 were induction failures, 8 in first chemotherapy-refractory relapse, 7 in untreated relapse, 3 in second or subsequent relapse, 4 were in second complete remission (CR), and 3 in second CR without platelet recovery (CRp), 2 were in high-risk CR1. Finally, 1 patient was in first CRp. Graft-versus-host disease (GVHD) prophylaxis was tacrolimus and mini-methorexate (MTX), and those who had an unrelated or 1 antigen-mismatched donor received low-dose rabbit-ATG (Thymoglobulin™). All patients engrafted. Forty-one patients had active leukemia at the time of transplant, and 35 achieved CR (85%). Twenty of the 42 AML patients and 5 of 9 CML patients are alive with a projected median overall survival (OS) of 23 months. Marrow and blood (T cell) chimerism studies at day +100 revealed that both in the lower-dose Clo groups (groups 1+2) and the higher-dose Clo groups (groups 3+4), the patients had a median of 100% donor (T cell)-derived DNA. There has been no secondary graft failure. In the first 100 days, 1 patient died of pneumonia, and 1 of liver GVHD. We conclude that (1) Clo ± Flu with i.v. Bu as pretransplant conditioning is safe in high-risk myeloid leukemia patients; (2) clofarabine is sufficiently immunosuppressive to support allo-SCT in myeloid leukemia; and (3) the median OS of 23 months in this high-risk patient population is encouraging. Additional studies to evaluate the antileukemic efficacy of Clo ± Flu with i.v. Bu as pretransplant conditioning therapy are warranted

2: Donor-Recipient Host-Versus-Graft Human Leukocyte Antigen Mismatches and Outcome of Cord Blood Transplants

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