Autophagy is Involved in Cardiac Remodeling in Response to Environmental Temperature Change.

Objectives: To study the reversibility of cold-induced cardiac hypertrophy and the role of autophagy in this process. Background: Chronic exposure to cold is known to cause cardiac hypertrophy independent of blood pressure elevation. The reversibility of this process and the molecular mechanisms involved are unknown. Methods: Studies were performed in two-month-old mice exposed to cold (4°C) for 24 h or 10 days. After exposure, the animals were returned to room temperature (21°C) for 24 h or 1 week. Results: We found that chronic cold exposure significantly increased the heart weight/tibia length (HW/TL) ratio, the mean area of cardiomyocytes, and the expression of hypertrophy markers, but significantly decreased the expression of genes involved in fatty acid oxidation. Echocardiographic measurements confirmed hypertrophy development after chronic cold exposure. One week of deacclimation for cold-exposed mice fully reverted the morphological, functional, and gene expression indicators of cardiac hypertrophy. Experiments involving injection of leupeptin at 1 h before sacrifice (to block autophagic flux) indicated that cardiac autophagy was repressed under cold exposure and re-activated during the first 24 h after mice were returned to room temperature. Pharmacological blockage of autophagy for 1 week using chloroquine in mice subjected to deacclimation from cold significantly inhibited the reversion of cardiac hypertrophy. Conclusion: Our data indicate that mice exposed to cold develop a marked cardiac hypertrophy that is reversed after 1 week of deacclimation. We propose that autophagy is a major mechanism underlying the heart remodeling seen in response to cold exposure and its posterior reversion after deacclimation.

Measuring interoception: The phase adjustment task.

Interoception, perception of one's bodily state, has been associated with mental health and socio-emotional processes. However, several interoception tasks are of questionable validity, meaning associations between interoception and other variables require confirmation with new measures. Here we describe the novel, smartphone-based Phase Adjustment Task (PAT). Tones are presented at the participant's heart rate, but out of phase with heartbeats. Participants adjust the phase relationship between tones and heartbeats until they are synchronous. Data from 124 participants indicates variance in performance across participants which is not affected by physiological or strategic confounds. Associations between interoception and anxiety, depression and stress were not significant. Weak associations between interoception and mental health variables may be a consequence of testing a non-clinical sample. A second study revealed PAT performance to be moderately stable over one week, consistent with state effects on interoception.

Autophagic control of cardiac steatosis through FGF21 in obesity-associated cardiomyopathy.

OBJECTIVE: High-fat diet-induced obesity leads to the development of hypertrophy and heart failure through poorly understood molecular mechanisms. We have recently shown that fibroblast growth factor-21 (FGF21) is produced by the heart and exerts protective effects that prevent cardiac hypertrophy development and oxidative stress. The aim of this study was to determine the effects of FGF21 on the cardiomyopathy associated with obesity development. RESULTS: Fgf21-/- mice showed an enhanced increase in the heart weight/tibia length (HW/TL) ratio in response to the high-fat diet. In keeping with this, echocardiographic measurements confirmed enhanced cardiac hypertrophy in Fgf21-/- mice. At the cellular level, the area of cardiomyocytes was increased in Fgf21-/- mice fed a high-fat diet. Furthermore, a high-fat diet induced fatty acid oxidation in the hearts of Fgf21-/- mice accompanied by an increase in cardiac oxidative stress. Oil-red O staining revealed the presence of higher amounts of lipid droplets in the hearts of Fgf21-/- mice fed a high-fat diet relative to wt mice fed this same diet. Finally, Fgf21-/- mice fed a high-fat diet showed impaired cardiac autophagy and signs of inactive cardiac lipophagy, suggesting that FGF21 promotes autophagy in cardiomyocytes. CONCLUSIONS: Our data indicate that a lack of FGF21 enhances the susceptibility of mice to the development of obesity-related cardiomyopathy. Furthermore, we demonstrate that this cardiac dysfunction is associated with deleterious lipid accumulation in the heart. An impaired ability of FGF21 to promote autophagy/lipophagy may contribute to lipid accumulation and cardiac derangements.

Protein phosphatase 1 regulatory subunit 1A in ewing sarcoma tumorigenesis and metastasis.

Protein phosphatase inhibitors are often considered as tumor promoters. Protein phosphatase 1 regulatory subunit 1A (PPP1R1A) is a potent protein phosphatase 1 (PP1) inhibitor; however, its role in tumor development is largely undefined. Here we characterize, for the first time, the functions of PPP1R1A in Ewing sarcoma (ES) pathogenesis. We found that PPP1R1A is one of the top ranked target genes of EWS/FLI, the master regulator of ES, and is upregulated by EWS/FLI via a GGAA microsatellite enhancer element. Depletion of PPP1R1A resulted in a significant decrease in oncogenic transformation and cell migration in vitro as well as xenograft tumor growth and metastasis in an orthotopic mouse model. RNA-sequencing and functional annotation analyses revealed that PPP1R1A regulates genes associated with various cellular functions including cell junction, adhesion and neurogenesis. Interestingly, we found a significant overlap of PPP1R1A-regulated gene set with that of ZEB2 and EWS, which regulates metastasis and neuronal differentiation in ES, respectively. Further studies for characterization of the molecular mechanisms revealed that activation of PPP1R1A by PKA phosphorylation at Thr35, and subsequent PP1 binding and inhibition, was required for PPP1R1A-mediated tumorigenesis and metastasis, likely by increasing the phosphorylation levels of various PP1 substrates. Furthermore, we found that a PKA inhibitor impaired ES cell proliferation, tumor growth and metastasis, which was rescued by the constitutively active PPP1R1A. Together, these results offered new insights into the role and mechanism of PPP1R1A in tumor development and identified an important kinase and phosphatase pathway, PKA/PPP1R1A/PP1, in ES pathogenesis. Our findings strongly suggest a potential therapeutic value of inhibition of the PKA/PPP1R1A/PP1 pathway in the treatment of primary and metastatic ES.

Rapid defense responses in maize leaves induced by Spodoptera exigua caterpillar feeding.

Insects such as the beet armyworm (Spodoptera exigua) cause extensive damage to maize (Zea mays). Maize plants respond by triggering defense signaling, changes in gene expression, and biosynthesis of specialized metabolites. Leaves of maize inbred line B73, which has an available genome sequence, were infested with S. exigua for 1 to 24 h, followed by comparisons of the transcript and metabolite profiles with those of uninfested controls. The most extensive gene expression responses occurred rapidly, within 4-6 h after caterpillar infestation. However, both gene expression and metabolite profiles were altered within 1 h and continued to change during the entire 24 h experiment. The defensive functions of three caterpillar-induced genes were examined using available Dissociation transposon insertions in maize inbred line W22. Whereas mutations in the benzoxazinoid biosynthesis pathway (Bx1 and Bx2) significantly improved caterpillar growth, the knockout of a 13-lipoxygenase (Lox8) involved in jasmonic acid biosynthesis did not. Interestingly, 9-lipoxygenases, which lead to the production of maize death acids, were more strongly induced by caterpillar feeding than 13-lipoxygenases, suggesting an as yet unknown function in maize defense against herbivory. Together, these results provide a comprehensive view of the dynamic transcriptomic and metabolomic responses of maize leaves to caterpillar feeding.

GvHD after umbilical cord blood transplantation for acute leukemia: an analysis of risk factors and effect on outcomes.

Using the Center for International Blood and Marrow Transplant Research (CIBMTR) registry, we analyzed 1404 umbilical cord blood transplantation (UCBT) patients (single (<18 years)=810, double (⩾18 years)=594) with acute leukemia to define the incidence of acute GvHD (aGvHD) and chronic GvHD (cGvHD), analyze clinical risk factors and investigate outcomes. After single UCBT, 100-day incidence of grade II-IV aGvHD was 39% (95% confidence interval (CI), 36-43%), grade III-IV aGvHD was 18% (95% CI, 15-20%) and 1-year cGvHD was 27% (95% CI, 24-30%). After double UCBT, 100-day incidence of grade II-IV aGvHD was 45% (95% CI, 41-49%), grade III-IV aGvHD was 22% (95% CI, 19-26%) and 1-year cGvHD was 26% (95% CI, 22-29%). For single UCBT, multivariate analysis showed that absence of antithymocyte globulin (ATG) was associated with aGvHD, whereas prior aGvHD was associated with cGvHD. For double UCBT, absence of ATG and myeloablative conditioning were associated with aGvHD, whereas prior aGvHD predicted for cGvHD. Grade III-IV aGvHD led to worse survival, whereas cGvHD had no significant effect on disease-free or overall survival. GvHD is prevalent after UCBT with severe aGvHD leading to higher mortality. Future research in UCBT should prioritize prevention of GvHD.

First Attempt of Sequential Living Donor Liver and Hematopoietic Stem Cell Transplantation in a Child With Advanced Hepatocellular Carcinoma: Case Report.

Effective therapeutic options for advanced hepatocellular carcinoma are limited. Hematopoietic stem cell transplantation may offer a graft-versus-tumor effect. Combined liver and hematopoietic stem cell transplantation from the same donor with preparatory conditioning may promote tolerogenicity to the liver allograft and offers the potential for immunosuppression withdrawal. We report our experience with the use of this approach in a pediatric patient with invasive hepatocellular carcinoma and pulmonary metastases who underwent a living-donor liver transplantation followed by reduced-toxicity myeloablative conditioning and hematopoietic stem cell transplant from the same parental donor. Neutrophil engraftment and full donor chimerism was achieved without liver allograft dysfunction. Despite normal liver function and marrow engraftment, the patient succumbed to multisystem organ failure from disseminated toxoplasmosis. At autopsy, there was no histologic evidence of tumor recurrence. No pulmonary nodules were found. Regardless of the unfortunate overall result, this case demonstrates preliminary feasibility of sequential living-donor liver transplantation and hematopoietic stem cell transplantation for unresectable and metastasized hepatic tumors. Future studies in select pediatric patients require evaluation of the optimal conditioning regimen and prevention strategies for opportunistic infections to determine both graft-versus-tumor effect on hepatic tumors and durability of tolerogenicity and possible immunosuppression withdrawal.

Thermogenic activation represses autophagy in brown adipose tissue.

BACKGROUND: Brown adipose tissue (BAT) thermogenesis is an adaptive process, essential for energy expenditure and involved in the control of obesity. Obesity is associated with abnormally increased autophagy in white adipose tissue. Autophagy has been proposed as relevant for brown-vs-white adipocyte differentiation; however, its role in the response of BAT to thermogenic activation is unknown. METHODS: The effects of thermogenic activation on autophagy in BAT were analyzed in vivo by exposing mice to 24 h cold condition. The effects of norepinephrine (NE), cAMP and modulators of lysosomal activity were determined in differentiated brown adipocytes in the primary culture. Transcript expression was quantified by real-time PCR, and specific proteins were determined by immunoblot. Transmission electron microscopy, as well as confocal microscopy analysis after incubation with specific antibodies or reagents coupled to fluorescent emission, were performed in BAT and cultured brown adipocytes, respectively. RESULTS: Autophagy is repressed in association with cold-induced thermogenic activation of BAT in mice. This effect was mimicked by NE action in brown adipocytes, acting mainly through a cAMP-dependent protein kinase A pathway. Inhibition of autophagy in brown adipocytes leads to an increase in UCP1 protein and uncoupled respiration, suggesting a repressing role for autophagy in relation to the activity of BAT thermogenic machinery. Under basal conditions, brown adipocytes show signs of active lipophagy, which is suppressed by a cAMP-mediated thermogenic stimulus. CONCLUSIONS: Our results show a noradrenergic-mediated inverse relationship between autophagy and thermogenic activity in BAT and point toward autophagy repression as a component of brown adipocyte adaptive mechanisms to activate thermogenesis.

Cellular engineering and therapy in combination with cord blood allografting in pediatric recipients.

Cord blood (CB) transplantation is an alternate source of human hematopoietic progenitor cells for allogeneic stem cell transplantation in children and adolescents with both malignant and nonmalignant diseases. Current limitations included delay in hematopoietic reconstitution, increased incidence of primary graft failure and slow cellular immunoreconstitution. These limitations lead to a significant increase in primary graft failure, infectious complications and increased transplant-related mortality. There is a number of experimental approaches currently under investigation including cellular engineering to circumvent these limitations. In this review, we summarize the recent findings of utilizing ex vivo CB expansion with Notch1 ligand Delta 1, mesenchymal progenitor cells, the use of human placenta-derived stem cells and CB-derived natural killer cells. Early and preliminary results suggest some of these experimental cellular strategies may in part ameliorate the incidence of primary graft failure, delays in hematopoietic reconstitution and/or slowness in cellular immune reconstitution following unrelated CB transplantation.

A prognostic model predicting autologous transplantation outcomes in children, adolescents and young adults with Hodgkin lymphoma.

Autologous hematopoietic cell transplantation (AutoHCT) is a potentially curative treatment modality for relapsed/refractory Hodgkin lymphoma (HL). However, no large studies have evaluated pretransplant factors predictive of outcomes of AutoHCT in children, adolescents and young adults (CAYA, age <30 years). In a retrospective study, we analyzed 606 CAYA patients (median age 23 years) with relapsed/refractory HL who underwent AutoHCT between 1995 and 2010. The probabilities of PFS at 1, 5 and 10 years were 66% (95% confidence interval (CI): 62-70), 52% (95% CI: 48-57) and 47% (95% CI: 42-51), respectively. Multivariate analysis for PFS demonstrated that at the time of AutoHCT patients with Karnofsky/Lansky score ⩾90, no extranodal involvement and chemosensitive disease had significantly improved PFS. Patients with time from diagnosis to first relapse of <1 year had a significantly inferior PFS. A prognostic model for PFS was developed that stratified patients into low-, intermediate- and high-risk groups, predicting for 5-year PFS probabilities of 72% (95% CI: 64-80), 53% (95% CI: 47-59) and 23% (95% CI: 9-36), respectively. This large study identifies a group of CAYA patients with relapsed/refractory HL who are at high risk of progression after AutoHCT. Such patients should be targeted for novel therapeutic and/or maintenance approaches post-AutoHCT.

Alternative donors extend transplantation for patients with lymphoma who lack an HLA matched donor.

Alternative donor transplantation is increasingly used for high-risk lymphoma patients. We analyzed 1593 transplant recipients (2000-2010) and compared transplant outcomes in recipients of 8/8 allele HLA-A, -B, -C and DRB1 matched unrelated donors (MUDs; n=1176), 7/8 allele HLA mismatched unrelated donors (MMUDs; n=275) and umbilical cord blood donors (1 or 2 units UCB; n=142). Adjusted 3-year non-relapse mortality of MMUD (44%) was higher as compared with MUD (35%; P=0.004), but similar to UCB recipients (37%; P=0.19), although UCB had lower rates of neutrophil and platelet recovery compared with unrelated donor groups. With a median follow-up of 55 months, 3-year adjusted cumulative incidence of relapse was lower after MMUD compared with MUD (25% vs 33%, P=0.003) but similar between UCB and MUD (30% vs 33%; P=0.48). In multivariate analysis, UCB recipients had lower risks of acute and chronic GVHD compared with adult donor groups (UCB vs MUD: hazard ratio (HR)=0.68, P=0.05; HR=0.35; P<0.001). Adjusted 3-year OS was comparable (43% MUD, 37% MMUD and 41% UCB). These data highlight the observation that patients with lymphoma have acceptable survival after alternative donor transplantation. MMUD and UCB can extend the curative potential of allotransplant to patients who lack suitable HLA matched sibling or MUD.

Sequential myeloablative autologous stem cell transplantation and reduced intensity allogeneic hematopoietic cell transplantation is safe and feasible in children, adolescents and young adults with poor-risk refractory or recurrent Hodgkin and non-Hodgkin lymphoma.

The outcome of children, adolescents and young adults (CAYA) with poor-risk recurrent/refractory lymphoma is dismal (⩽30%). To overcome this poor prognosis, we designed an approach to maximize an allogeneic graft vs lymphoma effect in the setting of low disease burden. We conducted a multi-center prospective study of myeloablative conditioning (MAC) and autologous stem cell transplantation (AutoSCT), followed by a reduced intensity conditioning (RIC) and allogeneic hematopoietic cell transplantation (AlloHCT) in CAYA, with poor-risk refractory or recurrent lymphoma. Conditioning for MAC AutoSCT consisted of carmustine/etoposide/cyclophosphamide, RIC consisted of busulfan/fludarabine. Thirty patients, 16 Hodgkin lymphoma (HL) and 14 non-Hodgkin lymphoma (NHL), with a median age of 16 years and median follow-up of 5years, were enrolled. Twenty-three patients completed both MAC AutoSCT and RIC AlloHCT. Allogeneic donor sources included unrelated cord blood (n=9), unrelated donor (n=8) and matched siblings (n=6). The incidence of transplant-related mortality following RIC AlloHCT was only 12%. In patients with HL and NHL, 10 year EFS was 59.8% and 70% (P=0.613), respectively. In summary, this approach is safe, and long-term EFS with this approach is encouraging considering the poor-risk patient characteristics and the use of unrelated donors for RIC AlloHCT in the majority of cases.

New frontiers in pediatric Allo-SCT.

The inaugural meeting of 'New Frontiers in Pediatric Allogeneic Stem Cell Transplantation' organized by the Pediatric Blood and Transplant Consortium (PBMTC) was held at the American Society of Pediatric Hematology and Oncology Annual Meeting. This meeting provided an international platform for physicians and investigators active in the research and utilization of pediatric Allo-SCT in children and adolescents with malignant and non-malignant disease (NMD), to share information and develop future collaborative strategies. The primary objectives of the conference included: (1) to present advances in Allo-SCT in pediatric ALL and novel pre and post-transplant immunotherapy; (2) to highlight new strategies in alternative allogeneic stem cell donor sources for children and adolescents with non-malignant hematological disorders; (3) to discuss timing of immune reconstitution after Allo-SCT and methods of facilitating more rapid recovery of immunity; (4) to identify strategies of utilizing Allo-SCT in pediatric myeloproliferative disorders; (5) to develop diagnostic and therapeutic approaches to hematological complications post pediatric Allo-SCT; (6) to enhance the understanding of new novel cellular therapeutic approaches to pediatric malignant and non-malignant hematological disorders; and (7) to discuss optimizing drug therapy in pediatric recipients of Allo-SCT. This paper will provide a brief overview of the conference.

Reduced toxicity, myeloablative conditioning with BU, fludarabine, alemtuzumab and SCT from sibling donors in children with sickle cell disease.

BU and CY (BU/CY; 200 mg/kg) before HLA-matched sibling allo-SCT in children with sickle cell disease (SCD) is associated with ~85% EFS but is limited by the acute and late effects of BU/CY myeloablative conditioning. Alternatives include reduced toxicity but more immunosuppressive conditioning. We investigated in a prospective single institutional study, the safety and efficacy of a reduced-toxicity conditioning (RTC) regimen of BU 12.8-16 mg/kg, fludarabine 180 mg/m(2), alemtuzumab 54 mg/m(2) (BFA) before HLA-matched sibling donor transplantation in pediatric recipients with symptomatic SCD. Eighteen patients, median age 8.9 years (2.3-20.2), M/F 15/3, 15 sibling BM and 3 sibling cord blood (CB) were transplanted. Mean whole blood and erythroid donor chimerism was 91% and 88%, at days +100 and +365, respectively. Probability of grade II-IV acute GVHD was 17%. Two-year EFS and OS were both 100%. Neurological, pulmonary and cardiovascular function were stable or improved at 2 years. BFA RTC and HLA-matched sibling BM and CB allo-SCT in pediatric recipients result in excellent EFS, long-term donor chimerism, low incidence of GVHD and stable/improved organ function.

Impact of telemonitoring home care patients with heart failure or chronic lung disease from primary care on healthcare resource use (the TELBIL study randomised controlled trial).

BACKGROUND: There is growing evidence that home telemonitoring can be advantageous in societies with increasing prevalence of chronic diseases.The main objective of this study is to evaluate the effect of a primary care-based telemonitoring intervention on the number and length of hospital admissions. METHODS: A randomised controlled trial was carried out across 20 health centres in Bilbao (Basque Country, Spain) to assess the impact of home telemonitoring on in-home chronic patients compared with standard care. The study lasted for one year. Fifty-eight in-home patients, diagnosed with heart failure (HF) and/or chronic lung disease (CLD), aged 14 or above and with two or more hospital admissions in the previous year were recruited. The intervention consisted of daily patient self-measurements of respiratory-rate, heart-rate, blood pressure, oxygen saturation, weight, body temperature and the completion of a health status questionnaire using PDAs. Alerts were generated when pre-established thresholds were crossed. The control group (CG) received usual care. The primary outcome measure was the number of hospital admissions that occurred at 12 months post-randomisation. The impact of telemonitoring on the length of hospital stay, use of other healthcare resources and mortality was also explored. RESULTS: The intervention group (IG) included 28 patients and the CG 30. Patient baseline characteristics were similar in both groups. Of the 21 intervention patients followed-up for a year, 12 had some admissions (57.1%), compared to 19 of 22 controls (86.4%), being the difference statistically significant (p = 0.033, RR 0.66; 95%CI 0.44 to 0.99). The mean hospital stay was overall 9 days (SD 4.3) in the IG versus 10.7 (SD 11.2) among controls, and for cause-specific admissions 9 (SD 4.5) vs. 11.2 (SD 11.8) days, both without statistical significance (p = 0.891 and 0.927, respectively). Four patients need to be telemonitored for a year to prevent one admission (NNT). There were more telephone contacts in the IG than in the CG (22.6 -SD 16.1- vs. 8.6 -SD 7.2-, p = 0.001), but fewer home nursing visits (15.3 -SD 11.6- vs. 25.4 -SD 26.3-, respectively), though the difference was not statistically significant (p = 0.3603). CONCLUSIONS: This study shows that telemonitoring of in-home patients with HF and/or CLD notably increases the percentage of patients with no hospital admissions and indicates a trend to reduce total and cause-specific hospitalisations and hospital stay. Home telemonitoring can constitute a beneficial alternative mode of healthcare provision for medically unstable elderly patients. TRIAL REGISTRATION: Current Controlled Trials ISRCTN89041993.

Allogeneic hematopoietic cell transplantation for neuroblastoma: the CIBMTR experience.

Although the role of autologous hematopoietic cell transplantation (auto-HCT) is well established in neuroblastoma (NBL), the role of allogeneic HCT (allo-HCT) is controversial. The Center for International Blood and Marrow Transplant Research conducted a retrospective review of 143 allo-HCT for NBL reported in 1990-2007. Patients were categorized into two different groups: those who had not (Group 1) and had (Group 2) undergone a prior auto-HCT (n=46 and 97, respectively). One-year and five-year OS were 59% and 29% for Group 1 and 50% and 7% for Group 2, respectively. Among donor types, disease-free survival (DFS) and OS were significantly lower for unrelated transplants at 1 and 3 years but not at 5 years post HCT. Patients in CR or very good partial response (VGPR) at transplant had lower relapse rates and better DFS and OS, compared with those not in CR or VGPR. Our analysis indicates that allo-HCT can cure some neuroblastoma patients, with lower relapse rates and improved survival in patients without a history of prior auto-HCT as compared with those patients who had previously undergone auto-HCT. Although the data do not address why either strategy was chosen for patients, allo-HCT after a prior auto-HCT appears to offer minimal benefit. Disease recurrence remains the most common cause of treatment failure.

Rescuing the neonatal brain from hypoxic injury with autologous cord blood.

Brain injury resulting from perinatal hypoxic-ischemic encephalopathy (HIE) is a major cause of acute mortality in infants and chronic neurologic disability in surviving children. Recent multicenter clinical trials demonstrated the effectiveness of hypothermia initiated within the first 6 postnatal hours to reduce the risk of death or major neurological disabilities among neonates with HIE. However, in these trials, approximately 40% of cooled infants died or survived with significant impairments. Therefore, adjunct therapies are required to improve the outcome in neonates with HIE. Cord blood (CB) is a rich source of stem cells. Administration of human CB cells in animal models of HIE has generally resulted in improved outcomes and multiple mechanisms have been suggested including anti-inflammation, release of neurotrophic factors and stimulation of endogenous neurogenesis. Investigators at Duke are conducting studies of autologous CB infusion in neonates with HIE and in children with cerebral palsy. These pilot studies indicate no added risk from the regimens used, but results of ongoing placebo-controlled trials are needed to assess efficacy. Meanwhile, further investigations are warranted to determine the best strategies, that is, timing, dosing, route of delivery, choice of stem cells and ex vivo modulations, to attain long-term benefits of CB stem cell therapy.