Search

Although emergency departments (EDs) play an integral role in the delivery of acute unscheduled care, they have not been fully integrated into broader health care reform efforts. Communication and coordination with the ambulatory environment remain limited, leaving ED care disconnected from patients' longitudinal care. In a value-based environment focused on improving quality, decreasing costs, enhancing population health, and improving the patient experience, this oversight represents a missed opportunity for emergency care. When integrated with primary and subspecialty care, emergency care might meet the needs of patients, providers, and payers more efficiently than yet realized. This article uses the Merit-Based Incentive Payment System from the Medicare Access and CHIP Reauthorization Act as a framework to outline a strategy for improving the value of emergency care, including integrating quality and resource use measures across health care delivery settings and populations, encouraging care coordination from the ED, and implementing robust health information exchange systems., (Copyright © 2016 American College of Emergency Physicians. Published by Elsevier Inc. All rights reserved.)

Background & Aims: Patients with anorexia nervosa (AN) have low serum IGF-I levels that may contribute to a lower bone mineral mass. We investigated the effects of a fermented, protein-fortified, dairy product on serum IGF-I levels in patients with AN during an in-hospital refeeding program., Methods: In this multicenter, randomized, double-blind, placebo-controlled, clinical trial conducted at 3 university hospitals and 3 private clinics in France and Switzerland, 62 women recently admitted with confirmed AN and with a baseline low serum IGF-I level were randomized to 2 daily isocaloric fresh cheese pots containing either 15 g/150 g or 3 g/150 g (controls) of protein for 4 weeks. The primary outcome was the change in IGF-I levels., Results: In the primary intention-to-treat analysis, mean serum IGF-I levels increased during the intervention phase from 22.9 ± 1.5 to 28.6 ± 1.3 nmol/L (means ± SEM) (+20.2%) in the intervention group and from 20.2 ± 1.2 to 25.7 ± 1.5 nmol/L (+16.8%) in controls. In a preplanned analysis of covariance with repeated measures, the between-group difference was close to statistical significance (P = 0.071). In a post-hoc mixed-regression model analysis, the difference was statistically significant (4.9 nmol/l increase; P = 0.003), as was the change of the ratio IGF-I/IGF-BP3 (P=0.004). There was no between-group difference in biochemical markers of bone turnover (osteocalcin, P1NP, CTX) or in serum parathyroid hormone level. Serum calcium levels slightly increased during the intervention phase in the higher protein group (P = 0.02). IGF-BP2 decreased significantly more in the intervention group during the follow up period at week 4 after supplements cessation (P = 0.019)., Conclusions: Intake of a fermented, protein-fortified, isocaloric dairy product during 4 weeks may slightly increase serum IGF-I levels in women with AN, without significant changes in bone turnover markers., Clinical Trial Registration Number: NCT01823822 (www.clinicaltrials.gov)., (Copyright © 2015 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.)

Background and Objective: Anorexia nervosa (AN) is associated with parameters involved in body mass index (BMI) regulation. Contrary to obesity, BMI kinetics around the adiposity rebound is not documented in AN. This study aimed at investigating which characteristics of BMI kinetics around the adiposity rebound are associated with AN., Methods: Multicentre case-control study with 101 inpatient women with AN onset after 10 years of age, and 101 healthy women, all free of overweight history and matched for age, level of education and fathers' socio-professional status. Age at adiposity rebound, pre- and post-adiposity rebound BMI velocities and accelerations (change in velocity over time) were estimated with linear mixed models using data recorded between 2 and 10 years of age., Results: Patients had an earlier adiposity rebound (mean (standard deviation (SD)): 5.3 (1.3) vs 5.7 (1.1) years), a larger BMI at adiposity rebound (mean (SD): 15.3 [1] vs 14.9 (0.9) kg/m 2 ) and 29% lower BMI acceleration after adiposity rebound than controls. After adjustment, only BMI at adiposity rebound and BMI acceleration after adiposity rebound were associated with a higher risk of AN (Odds ratio [95% confidence interval]: 2.15 [1.41-3.46] for an increase of 1 kg/m 2 and 2.44 [1.56-4.02] for an increase of 0.1 kg/(m 2 *years 2 ) respectively). These two factors were not correlated in patients (r = 0.007, p = 0.96)., Conclusions: A flattened evolution of BMI after adiposity rebound and higher BMI at adiposity rebound were associated with AN. Further prospective study is needed to confirm these findings., (Copyright © 2016 European Society for Clinical Nutrition and Metabolism. Published by Elsevier Ltd. All rights reserved.)

Binge eating has been usually viewed as a loss of control and an impulsive behavior. But, little is known about the actual behavior of binging patients (prevalently women) in terms of basic decision-making under risk or under uncertainty. In healthy women, stressful cues bias behavior for safer options, raising the question of whether food cues that are perceived as threatening by binging patients may modulate patients' behaviors towards safer options. A cross-sectional study was conducted with binging patients (20 bulimia nervosa (BN) and 23 anorexia nervosa binging (ANB) patients) and two control groups (22 non-binging restrictive (ANR) anorexia nervosa patients and 20 healthy participants), without any concomitant impulsive disorder. We assessed decisions under risk with a gambling task with known probabilities and decisions under uncertainty with the balloon analog risk taking task (BART) with unknown probabilities of winning, in three cued-conditions including neutral, binge food and stressful cues. In the gambling task, binging and ANR patients adopted similar safer attitudes and coherently elicited a higher aversion to losses when primed by food as compared to neutral cues. This held true for BN and ANR patients in the BART. After controlling for anxiety level, these safer attitudes in the food condition were similar to the ones under stress. In the BART, ANB patients exhibited a higher variability in their choices in the food compared to neutral condition. This higher variability was associated with higher difficulties to discard irrelevant information. All these results suggest that decision-making under risk and under uncertainty is not fundamentally altered in all these patients.

Objective: The role of planning in binge eating episodes is unknown. We investigated the characteristics of planning associated with food cues in binging patients. We studied planning based on backward reasoning, reasoning that determines a sequence of actions back to front from the final outcome., Method: A cross-sectional study was conducted with 20 healthy participants, 20 bulimia nervosa (BN), 22 restrictive (ANR) and 23 binging anorexia nervosa (ANB), without any concomitant impulsive disorder. In neutral/relaxing, binge food and stressful conditions, backward reasoning was assessed with the Race game, promotion of delayed large rewards with an intertemporal discounting task, attention with the Simon task, and repeating a dominant behavior with the Go/No-go task., Results: BN and to a lower extent ANB patients succeeded more at the Race game in food than in neutral condition. This difference discriminated binging from non-binging participants. Backward reasoning in the food condition was associated with lower approach behavior toward food in BN patients, and higher food avoidance in ANB patients. Enhanced backward reasoning in the food condition related to preferences for delayed large rewards in BN patients. In BN and ANB patients the enhanced success rate at the Race game in the food condition was associated with higher attention paid to binge food., Conclusion: These findings introduce a novel process underlying binges: planning based on backward reasoning is associated with binges. It likely aims to reduce craving for binge foods and extend binge refractory period in BN patients, and avoid binging in ANB patients. Shifts between these goals might explain shifts between eating disorder subtypes.

Objectives: To examine trends in out-of-pocket spending and the financial burden of care for persons with diabetes between 2001 and 2009, and to examine whether these trends are consistent with trends in access to prescription drugs and utilization of hospital services., Study Design and Methods: Data are from the 2001 to 2009 Medical Expenditure Panel Survey (MEPS). The sample includes persons aged 18 to 64 years with diagnosed diabetes. The primary outcome variable is the percent of people with out-of-pocket spending on insurance premiums and services that exceed 10% of family income. Secondary outcome measures include the percent with diabetes-related prescription drug use, perceived access to prescription drugs, hospital inpatient stays, and emergency department use in the past 12 months. Multiple regression analysis is used to control for changes in comorbid chronic conditions and other characteristics of persons with diabetes., Results: Both out-of-pocket spending and the percent with high financial burden decreased markedly for persons with diabetes between 2001 to 2003 and 2007 to 2009. The decrease in spending was driven primarily by a decrease in spending on prescription drugs, including diabetes-related prescriptions. The shift from brand name drugs to generics accounts for much of this decline, although decreases in out-of-pocket spending for both brand name and generic drugs also contributed. During the same period, utilization of and access to diabetes-related prescriptions increased, and hospital use decreased., Conclusions: Although the prevalence of diagnosed diabetes continues to increase, treatment is becoming more affordable, especially prescription drugs. This may offset some of the costs to the healthcare system of higher prevalence by reducing complications of uncontrolled diabetes that result in more costly hospital use.

Objectives: We used the 2003-2009 Medical Expenditure Panel Survey to evaluate average annual total and out-of-pocket expenditures by nonelderly adults with asthma., Study Design: We divided patients diagnosed with asthma into 4 groups, based on whether or not they had had an asthma attack in the previous year (a crude marker for disease severity) and whether or not they reported using treatment for their asthma., Methods: For each group we calculated total and out-of-pocket average annual spending for hospital inpatient, hospital outpatient, emergency department, and physician office care, as well as for prescription drugs. These averages were adjusted to account for differences in respondents' overall health (presence of other co-morbidities, self-reported health status, and self-reported activity limitations), sociodemographic characteristics (age, sex, race/ethnicity, income), and insurance status., Results: We found that among the 4 groups, those who were receiving treatment but continued to experience asthma attacks had the highest total and out-of-pocket expenditures in all categories, consistent with their likely higher illness severity. However, patients who reported receiving treatment and did not experience attacks also reported relatively high adjusted total and out-of-pocket expenditures-most notably $536 per year out of pocket for prescription medications and $231 per year out of pocket for physician office visits. After adjustment, about the same proportion of patients in these 2 groups (13.5% who did not get treated and had attacks, and 13.8% who did get treated and avoided attacks) reported high financial burden., Conclusions: Patients may experience financial challenges to appropriate self-management of asthma, even when they are able to avoid exacerbations.

Cryopreservation is a common procedure widely used in biological and clinical sciences. Similar protocols are also applied in preserving cancer stem cells, a field with high promises and challenges. Specific cell surface membrane proteins are considered to be biomarkers of cancer stem cells and they may play a critical role in differentiating stem cells from non stem cells. We have looked at the possible effect of long-term cryopreservation on the molecular integrity of breast MCF7 and lung, A549 and H460, cancer stem cells and to assess if these cells are more sensitive to long-term storage process. We analyzed the expression of CD24 and CD38 as two potent biomarkers of lung cancer stem cells and EpCAM and ALDH that are used as biomarkers of a wide range of cancer stem cells. We also selected three genes essential for the normal functioning of the cells, Fos, MUC1, and HLA. Our results indicate a pattern of down-regulation in the expression of the genes following freezing, in particular among cell surface marker proteins. Global gene expression of the post-thaw breast and lung cancer stem cells also reveals a significant down-regulation in freeze-thaw cells independent from each other. Analyzing the canonical pathways between two populations reveals a significant alteration in the gene expression of the pathways involved in cell cycle, mitosis, and ataxia telangiectasia mutated pathways. Overall, our results indicate that current protocols for long-term storage of lung and breast cancer stem cells may substantially influence the activity and function of genes.

Background: With growing pressure to improve the quality and coordination of care, physicians feel a need to streamline their relationships with other practitioners around shared care for patients. Some physicians have developed written agreements that articulate the respective responsibilities of 2 or more parties for coordination of patient care, ie, care coordination agreements (CCAs)., Objectives: To describe how CCAs are formed and explore facilitators and barriers to adoption of effective CCAs, the extent to which CCAs may be replicable in different market contexts, and the implications for policies and programs that aim to improve the coordination of care., Study Design: Qualitative study of primary care physicians participating in CCAs and representatives of their specialist, hospital, or community-based partners., Methods: Semi-structured interviews with participating providers and national thought leaders in care coordination were reviewed to develop key themes., Results: Agreements that address referral and access processes were considered useful by all practices that had implemented them. Practices that implemented agreements including guidance on shared management of specific clinical conditions (comanagement) also found them useful. CCAs were most successful in settings where both parties to the agreement already had stable communication pathways (such as an electronic health record [EHR], designated staff) and strong working relationships., Conclusions: Policy changes (such as shifts in reimbursement to favor collaborative care or clarification of laws governing such collaborations) can help to support the development and implementation of CCAs, and can address factors that currently make some markets less supportive of coordination.

Background: Despite expectations that medical homes provide "24 × 7 coverage" there is little to guide primary care practices in developing sustainable models for accessible and coordinated after-hours care., Objective: To identify and describe models of after-hours care in the U.S. that are delivered in primary care sites or coordinated with a patient's usual primary care provider., Design: Qualitative analysis of data from in-depth telephone interviews., Setting: Primary care practices in 16 states and the organizations they partner with to provide after-hours coverage., Participants: Forty-four primary care physicians, practice managers, nurses and health plan representatives from 28 organizations., Approach: Analyses examined after-hours care models, facilitators, barriers and lessons learned., Results: Based on 28 organizations interviewed, five broad models of after-hours care were identified, ranging in the extent to which they provide continuity and patient access. Key themes included: 1) The feasibility of a model varies for many reasons, including patient preferences and needs, the local health care market supply, and financial compensation; 2) A shared electronic health record and systematic notification procedures were extremely helpful in maintaining information continuity between providers; and 3) after-hours care is best implemented as part of a larger practice approach to access and continuity., Conclusion: After-hours care coordinated with a patient's usual primary care provider is facilitated by consideration of patient demand, provider capacity, a shared electronic health record, systematic notification procedures and a broader practice approach to improving primary care access and continuity. Payer support is important to increasing patients' access to after-hours care.

Being prepared for a natural disaster, infectious disease outbreak or other emergency where many injured or ill people need medical care while maintaining ongoing operations is a significant challenge for local health systems. Emergency preparedness requires coordination of diverse entities at the local, regional and national levels. Given the diversity of stakeholders, fragmentation of local health care systems and limited resources, developing and sustaining broad community coalitions focused on emergency preparedness is difficult. While some stakeholders, such as hospitals and local emergency medical services, consistently work together, other important groups--for example, primary care clinicians and nursing homes--typically do not participate in emergency-preparedness coalitions, according to a new qualitative study of 10 U.S. communities by the Center for Studying Health System Change (HSC). Challenges to developing and sustaining community coalitions may reflect the structure of preparedness activities, which are typically administered by designated staff in hospitals or large medical practices. There are two general approaches policy makers could consider to broaden participation in emergency-preparedness coalitions: providing incentives for more stakeholders to join existing coalitions or building preparedness into activities providers already are pursuing. Moreover, rather than defining and measuring processes associated with collaboration--such as coalition membership or development of certain planning documents--policy makers might consider defining the outcomes expected of a successful collaboration in the event of a disaster, without regard to the specific form that collaboration takes.

Contrary to conventional wisdom that Medicaid patients often use hospital emergency departments (EDs) for routine care, the majority of ED visits by nonelderly Medicaid patients are for symptoms suggesting urgent or more serious medical problems, according to a new national study by the Center for Studying Health System Change (HSC). About 10 percent of nonelderly Medicaid patient ED visits are for nonurgent symptoms, compared with about 7 percent for privately insured nonelderly people. Nonetheless, there are clearly opportunities to develop less-costly care options than emergency departments for both nonelderly Medicaid and privately insured patients. To reduce ED use, policy makers might consider how to encourage development of care settings that can quickly handle a high volume of potentially urgent medical problems. Policy makers may want to focus initially on conditions that account for high ED volume that could likely be treated in less resource-intensive settings. For example, diagnoses of acute respiratory and other common infections in children and injuries together account for about 53 percent of ED visits by children aged 0 to 12 covered by Medicaid and almost 60 percent of ED visits by privately insured children aged 0 to 12. While some infections and injuries will be too serious to treat elsewhere, lower-cost settings that can provide a moderate intensity of care and urgent response time likely could reduce emergency department use.

Previous research suggests that some substance users have multiple crisis detoxification visits and never access rehabilitation care. This care-seeking pattern leads to poorer outcomes and higher costs. The authors aimed to identify predictors of repeat detoxification visits by analyzing state-level data routinely collected at the time of substances use services admission. Repeat detoxification clients were more likely to be homeless, city-dwelling fee-for-service Medicaid recipients. Repeat detoxification clients were less likely than those with one admission to enter rehabilitation within 3 days. Treatment providers should aim for rapid transfer to rehabilitation and consider expanding detoxification intake data to improve risk stratification., (© Taylor & Francis Group, LLC)

Due to the recent advancements in stem cell biology and engineering, scientists have been increasingly interested in creating in vitro niches for embryonic and adult stem cells, and, following induction and differentiation with the appropriate media, the production of large scale blood production. This artificially created niche for hematopoietic cells will be composed of three materials: the stem cells themselves, the scaffold surrounding the stem cell, and the media used to expand and differentiate the stem cells. This paper will examine the recent advancements in technology for each of these relating to the development of an artificial stem cell niche. Many key aspects of the artificial niche need to be improved on before we can scale up the engineered device for large scale blood production including more efficient methods of retrieval of the embroid bodies produced from the microfluidic channels. The current state of experimental methods such as these as well as relevant discoveries in related fields that could be applied to artificial niche technology is described in this paper. Furthermore, we present a mathematical model to describe cell expansion in the artificial hematopoietic stem cell niche in order to design and optimize a scaled-up bioreactor. The mathematical model describes the dynamics of expansion, and maintenance of homeostasis in the bioreactor.

In 2006, the Institute of Medicine (IOM) advanced the concept of "coordinated, regionalized, and accountable emergency care systems" to address significant problems with the delivery of emergency medical care in the United States. Achieving this vision requires the thoughtful implementation of well-aligned, system-level structures and processes that enhance access to emergency care and improve patient outcomes at a sustainable cost. Currently, the delivery of emergency medical care is supported by numerous administrative systems, including economic; reimbursement; legal and regulatory structures; licensure, credentialing, and accreditation processes; medicolegal systems; and quality reporting mechanisms. In addition, many regionalized systems may not optimize patient outcomes because of current administrative barriers that make it difficult for providers to deliver the best care. However, certain administrative barriers may also threaten the sustainability of integration efforts or prevent them altogether. This article identifies significant administrative challenges to integrating networks of emergency care in four specific areas: reimbursement, medical-legal, quality reporting mechanisms, and regulatory aspects. The authors propose a research agenda for indentifying optimal approaches that support consistent access to quality emergency care with improved outcomes for patients, at a sustainable cost. Researching administrative challenges will involve careful examination of the numerous natural experiments in the recent past and will be crucial to understand the impact as we embark on a new era of health reform., (2010 by the Society for Academic Emergency Medicine.)

The ideal emergency care system delivers the right care to the right patient at the right time and yields appropriate patient outcomes at a sustainable overall cost. Transforming the current system of emergency care into the Institute of Medicine's vision of a coordinated, regionalized, and accountable emergency care system requires careful consideration of administrative challenges and barriers. Left unaddressed, certain processes, systems, and structures may prevent integration efforts or threaten long-term viability., (2010 by the Society for Academic Emergency Medicine.)

Recent years have brought significant breakthroughs in the understanding of tumor biology, related to discovery of cancer stem cells (CSCs) in acute myelogenous leukemia as well as in a number of solid tumors. This finding revealed that not all tumor cells are able to divide indefinitely, and that the bulk of tumor cells are expanded because of divisions and differentiation of CSC fraction. Although the CSCs identified in acute leukemia have a phenotype of early hematopoietic progenitors, it seems that CSCs in multiple myeloma (MM) may resemble the memory B cell fraction. Previous studies in patients with MM have documented the existence of cells without plasma cell characteristics expressing MM-type immunoglobulin genes--so-called "clonotypic" B cells. These cells have been characterized functionally and phenotypically as chemoresistant recirculating B cells. They have been found to self-renew and to be capable of initiating MM growth in immunocompromised animals. Controversy exists as to whether these cells truly belong to an MM clone, however; they may represent only the remaining clones of premalignant B cells. The identification of MM stem cells responsible for the recurrence of MM is of primary importance in designing targeted therapies to definitely cure this disease. This article summarizes the current state of knowledge on these hypothetical "MM stem cells.", (Copyright 2010 American Society for Blood and Marrow Transplantation. Published by Elsevier Inc. All rights reserved.)

Induction of tumor-specific immunity is an attractive approach to cancer therapy, however to date every major pivotal trial has resulted in failure. While the phenomena of tumor-mediated immune suppression has been known for decades, only recently have specific molecular pathways been elucidated, and for the first time, rationale means of intervening and observing results of intervention have been developed. In this review we describe major advances in our understanding of tumor escape from immunological pressure and provide some possible therapeutic scenarios for enhancement of efficacy in future cancer vaccine trials., (Copyright (c) 2010 Elsevier Inc. All rights reserved.)

Use of care management tools--such as group visits or patient registries--varies widely among primary care physicians whose practices care for patients with four common chronic conditions--asthma, diabetes, congestive heart failure and depression--according to a new national study by the Center for Studying Health System Change (HSC). For example, less than a third of these primary care physicians in 2008 reported their practices use nurse managers to coordinate care, and only four in 10 were in practices using registries to keep track of patients with chronic conditions. Physicians also used care management tools for patients with some chronic conditions but not others. Practice size and setting were strongly related to the likelihood that physicians used care management tools, with solo and smaller group practices least likely to use care management tools. The findings suggest that, along with experimenting with financial incentives for primary care physicians to adopt care management tools, policy makers might consider developing community-level care management resources, such as nurse managers, that could be shared among smaller physician practices.

"Niche" is defined as a specialized regulatory microenvironment, consisting of components which control the fate specification of stem and progenitor cells, as well as maintaining their development by supplying the requisite factors. Bone marrow (BM) niche has a well-organized architecture and is composed of osteoblasts, osteoclasts, bone marrow endothelial cells, stromal cells, adipocytes and extracellular matrix proteins (ECM). These elements play an essential role in the survival, growth and differentiation of diverse lineages of blood cells, but also provide optimal growth environment for multiple hematological malignancies including multiple myeloma (MM). MM is a neoplastic plasma cell disorder which not only resides in BM but also converts it into specialized neoplastic niche. This niche aids the growth and spreading of tumor cells by a complex interplay of cytokines, chemokines, proteolytic enzymes and adhesion molecules. Moreover, the MM BM microenvironment was shown to confer survival and chemoresistance of MM cells to current therapies. However, our knowledge in this field is still in infancy and many details are unknown. Therefore, there is a strong need to further dissect the MM BM niche and understand the process of how the complex interactions with BM milieu influence MM growth, survival and development of resistance to chemotherapy. A better and more detailed understanding of neoplastic MM niche will provide a guiding model for identifying and validating novel targeted therapies directed against MM. Therefore, in the present review, we have focused principally on the basic features, physical structures, and functions of the BM niche and have highlighted its interaction with MM cells.

Translation of small interfering RNA (siRNA)-based approaches into practical therapeutics is limited because of lack of an effective and cell-specific delivery system. Herein, we present a new method of selectively delivering siRNA to dendritic cells (DCs) in vivo using CD40 siRNA-containing immunoliposomes (siILs) that were decorated with DC-specific DEC-205 mAb. Administration of CD40 siILs resulted in DC-specific cell targeting in vitro and in vivo. On treatment with CD40 siILs, the expression of CD40 in DCs, as well allostimulatory activity was inhibited. In vivo administration resulted in selective siRNA uptake into immune organs and functional immune modulation as assessed using a model antigen. In conclusion, this is the first demonstration of DC-specific siRNA delivery and gene silencing in vivo, which highlights the potential of DC-mediated immune modulation and the feasibility of siRNA-based clinical therapy.

Animal studies have demonstrated that selective tropism of mesenchymal stem cells (MSC) for glioma may be used as a means of selective delivery of cytotoxic payloads. Endometrial Regenerative Cells (ERC) are a population of mesenchymal-like cells which possesse pluripotent differentiation capacity and is characterized by unique surface markers and growth factor production. In this study we sought to determine whether unmanipulated ERC would alter the growth of glioma using the aggressive C6/LacZ7 (C6) into Sprague Dawley rat model. ERC administration by intravenous (i.v.) or intratumoral (i.t.) showed significant inhibition of glioma: volume reduction of 49% after i.v. treatment (p < 0.05), and about 46% i.t. treatment (p < 0.05). Tumor reduction was associated with inhibition of angiogenesis and reduced numbers of CD133 positive cells in the incranial tumor. Despite the angiogenic potential of ERC in the hindlimb ischemia model, these data support a paradoxical tumor inhibitory activity of ERC. Further studies are needed to determine the qualitative differences between physiological angiogenesis, which seems to be supported by ERC and tumor angiogenesis which appeared to be inhibited.

Relapse of malignancy after allogeneic hematopoietic cell transplantation (allo-HCT) remains a therapeutic challenge. Blockade of the CTLA4 molecule can effectively augment antitumor immunity mediated by autologous effector T cells. We have assessed the safety and preliminary efficacy of a neutralizing, human anti-CTLA4 monoclonal antibody, ipilimumab, in stimulating the graft-versus-malignancy (GVM) effect after allo-HCT. Twenty-nine patients with malignancies that were recurrent or progressive after allo-HCT, received ipilimumab as a single infusion at dose cohorts between 0.1 and 3.0 mg/kg. Dose-limiting toxicity was not encountered, and ipilimumab did not induce graft-versus-host disease (GVHD) or graft rejection. Organ-specific immune adverse events (IAE) were seen in 4 patients (grade 3 arthritis, grade 2 hyperthyroidism, recurrent grade 4 pneumonitis). Three patients with lymphoid malignancy developed objective disease responses following ipilimumab: complete remission (CR) in 2 patients with Hodgkin disease and partial remission (PR) in a patient with refractory mantle cell lymphoma. At the 3.0 mg/kg dose, active serum concentrations of ipilimumab were maintained for more than 30 days after a single infusion. Ipilimumab, as administered in this clinical trial, does not induce or exacerbate clinical GVHD, but may cause organ-specific IAE and regression of malignancy. This study is registered at (http://clinicaltrials.gov) under NCI protocol ID P6082.

The mast cell-derived serine protease chymase is importantly involved not only in degradation, but in synthesis of bioactive peptides as well. Several studies suggest that chymase is the predominant enzyme in the production of angiotensin II (Ang II) from angiotensin-I in interstitial tissues. Interestingly, chymase has also been suggested to mature endothelin-1 (ET-1) from its precursor, big-ET-1 in vitro. The lack of availability of specific chymase inhibitors, beyond the chymotrypsin-like inhibitor chymostatin, currently hampers the investigation of the chymase/ET-1/Ang II paradigm in physiology and cardiovascular diseases. Nonetheless, the recent advent of highly selective chymase inhibitors is shedding new light on the role of this enzymatic pathway in the several inflammatory prone vascular diseases as summarized in the present review. Considering increasing evidence towards significant interactions between Ang II and ET-1 in cardiovascular diseases, the present review will address the role of chymase in the production of those two peptides. Whether chymase-dependent production of ET-1 plays an important role in cardiovascular pathologies will also be discussed.

To examine the role of endothelin ETA and ETB receptors in congestive heart failure due to cardiomyopathy, the effect of chronic treatment with selective ETA- and ETB-receptor antagonists (atrasentan and A-192621, respectively), alone and in combination, was assessed on functional and biochemical parameters of 52-week-old Bio 14.6 cardiomyopathic hamsters. Compared with control animals, cardiomyopathic hamsters treated for 9 weeks with atrasentan showed no variation in MAP; however, selective ETB- and combined nonselective ETA- and ETB-receptor antagonists increased systemic blood pressure. After selective ETB-receptor blockade, plasma endothelin levels were augmented. Importantly, this increase was highly enhanced (more than 8-fold) by concomitant ETA-receptor antagonism. Furthermore, the left ventricle:body weight ratio of cardiomyopathic hamsters treated with A-192621, alone or in combination with atrasentan, was significantly increased. On the other hand, decreased left ventricular end-diastolic pressure was observed in cardiomyopathic hamsters after selective ETA- or combined nonselective ETA/ETB-receptor antagonism, while only selective ETA-receptor blockade reduced left ventricular endothelin levels. Our results suggest that, in congestive heart failure, ETB receptors are essential to limit circulating endothelin levels, which may argue for improved cardiac benefits after long-term treatment with highly selective ETA-receptor antagonists.

Autologous or allogeneic SCT with conventional conditioning (chemotherapy with or without irradiation) has emerged as an effective and potentially curative therapy in patients with hematologic malignancies and in other selected solid tumors; however, several patients experience significant early and delayed side effects, including long-term endocrine imbalance and infertility. In spite of several reproductive recovery and pregnancy reports published in the oncology literature, review of medical literature reveals a paucity of comparable information in the SCT field. We report here four cases of ovarian recovery in patients who received hormonal replacement therapy after diagnosis of primary ovarian failure due to high-dose chemotherapy and SCT.

Background and Purpose: Calcitonin gene-related peptide (CGRP), a capsaicin-sensitive neuromodulator of splanchnic vascular tone in several animal species, remains poorly investigated in mouse models. We therefore assessed whether endogenous CGRP is a non-adrenergic/non-cholinergic (NANC) neuromodulator in the mesenteric vascular bed of the mouse., Experimental Approach: Arterial and venous changes in perfusion pressure in response to perivascular nerve stimulation (PNS) were monitored in the mouse mesenteric bed under basal conditions or precontracted with KCl (artery) or U46619 (vein) in circuits pretreated with guanethidine, atropine, indomethacin and prazosin. Arterial responses to NANC were also characterized with a CGRP1 antagonist, halphaCGRP8-37. Finally, the PNS-induced release of arterial CGRP was measured by enzyme immunoassay., Key Results: HalphaCGRP8-37 enhanced PNS-induced arterial increases in perfusion pressure under basal tone. PNS-induced stimulation of NANC triggered an halphaCGRP8-37 or capsaicin- sensitive reduction in perfusion pressure of the pre-contracted arterial bed only. Chemical removal of the endothelium inhibited PNS- and halphaCGRP- induced reduction in perfusion pressure in the arterial mesenteric bed. Responses to NANC nerves were reduced by guanylate and adenylate cyclase inhibitors (1H-[1,2,4]oxadiazole[4,3-a] quinoxalin-1-one (ODQ)) and [9-(tetrahydro-2-furanyl)-9H-purin-6-amine] (SQ 22,536), respectively. A neuronal NOS inhibitor (7-nitroindazole; 7-NI) also enhanced the response to NANC in vessels from wild-type, eNOS KO but not iNOS KO mice. Finally, PNS enhanced the release of immunoreactive CGRP from the perfused arterial mesenteric bed., Conclusions and Implications: Our study demonstrates a role for CGRP in the NANC-dependent reduction in perfusion pressure of the arterial but not venous mesenteric bed of the mouse.

Nitric oxide and other reactive oxygen species generated by nitric-oxide synthases (NOS) modulate, among several other cellular responses, the production of eicosanoids and platelet aggregation. The roles of specific NOS in these two phenomena remain to be determined. Thus, the present study assessed whether inducible NOS (iNOS) and endothelial NOS (eNOS) modulate in a similar manner the production of eicosanoids and platelet aggregation. Mice knocked out for eNOS (eNOS-/-) or iNOS (iNOS-/-) and their wild-type (WT) congeners were used to analyze agonist-induced increases in plasma levels of eicosanoids as well as inhibition of platelet aggregation ex vivo. Systemically administered endothelin-1 (ET-1) triggered an increase in plasma levels of 6-keto prostaglandin F(1alpha) (6-keto PGF(1alpha)) in WT and eNOS-/- but not in iNOS-/- mice. ET-1 (0.01-1 nmol/kg) also induced a dose-dependent inhibition of platelet aggregation in WT and eNOS-/- but not in iNOS-/- mice. Another agonist, bradykinin (10 nmol/kg), triggered the release of 6-keto PGF(1alpha) and inhibited platelet aggregation in all strains of mice studied. In addition, ADP-induced platelet aggregation in vitro was similarly reduced by iloprost (100 nM) in iNOS-/- mice and WT congeners. In another series of experiments, ET-1 (0.1 nmol/kg) significantly increased 8-isoprostane plasma levels in WT but not in iNOS-/- mice. Finally, a 3-week treatment with anti-oxidants inhibited the capacity of ET-1 to significantly increase plasma 6-keto PGF(1alpha) in WT mice. We show for the first time that iNOS is involved in the control of ET-1-induced prostacyclin release and related inhibition of platelet aggregation in the murine model.

The primary objective of this work is to determine the repairing potential of murine embryonic stem cells (ES) in murine model of Crohn's disease (CD). Colitis, induced in IL10-/- KO mice using piroxicam, was associated with the increased levels of IL-12. Enhanced yellow fluorescent protein (EYFP) marked murine ES cells (R1/129) and control non-fluorescent ES cells were subjected to in vitro differentiation into intestinal epithelial cells. IL 10-/- KO mice were injected with pre-differentiated ES-YFP cells and sacrificed after 2 and 3 months. Histopathological analysis of intestines demonstrated a progressive improvement in colitis (from grade-4 to grade-1 and -0) and decreased levels of IL-12 cytokine following transplantation. Fluorescent and confocal microscopy demonstrated presence of ES-EYFP cells in the colon, small intestine, liver, and thymus tissues but none in the spleen and bone marrow. The EYFP signal was not detected in sham (non-transplanted mice with induced colitis) and control IL10-/- KO mice. Engraftment, detected at 3 months post-transplant, correlated with markedly improved grading in colon histology (grade-1 or -0) and weight gain, as well as with decreased rectal prolapses. In vitro pre-differentiated ES cells migrated and homed exclusively into the colon, small intestine, and the liver, engrafted for long term, reduced inflammation and tissue damage, and restored immune balance. These findings suggest that pre-differentiated ES cells may become alternative source of stem cell therapy for CD with dual functions i.e. regenerating damaged epithelium and restoring immune imbalance occurring in this disease.

Cellular therapies derived from embryonic stem (ES) cells have gained a renewed interest with the experimental demonstration that an embryonic stem cell lines can be established from human blastocyst-stage embryos and prompted to differentiate into almost all types of cells present in the body including hematopoietic cells. Hematopoiesis is a series of cellular processes whereby short-lived mature blood cells are continuously replenished from a pool of rare pluripotential hematopoietic stem cells, in a highly orchestrated process. Aberrances in this intricate process may lead to a malignancy of essential blood-forming organs, causing diseases such as leukemia, aplastic anemia, lymphoma, myelodysplasia and myeloproliferative disorders. Embryonic stem cells show great potential and it may be technologically feasible to transplant differentiated ES cells and to cure various kinds of blood disorders. Understanding the biology of ES cell derived hematopoiesis may lead to the development of co-transplantation protocols that will result in a decreased morbidity and mortality by providing safer and simpler transplantation procedures for patients with malignant and non-malignant conditions. The potential utility of ES cells for gene therapy, tissue engineering and the treatment of a wide variety of currently untreatable diseases is simply too essential to ignore, however, our knowledge and ability to deliver these forms of therapy in a safe and efficient manner requires additional advances in the understanding of the basic biology of ES cells. In this article, we will discuss the factors and methodologies responsible for the differentiation of ES cells into hematopoietic progenitors and their potential to treat different blood related diseases.

The role of thrombopoietin (TPO) in adult hematopoiesis is well-established. A recent report suggests that TPO and vascular endothelial growth factor (VEGF) play a role in promoting formation of early erythropoietic progenitors in a nonhuman primate embryonic stem cell (ES) model. No such report exists for human ES cells as yet. Because TPO may become an important factor promoting human ES cell-derived hematopoiesis, we sought to investigate whether TPO in combination with VEGF can enhance human ES-derived hematopoiesis in an EB-derived culture system. The emphasis of this work was to demonstrate the molecular mechanisms involved in this process, specifically the role of c-mpl and its ligand TPO. Human ES cells were cultured to the EB state, and EB-derived secondary cultures supporting hematopoietic differentiation were established: condition 1, control (stem cell factor [SCF] and Flt3 ligand [Flt3L]); condition 2, SCF, Flt3L, and TPO; and condition 3, SCF, Flt3L, TPO, and VEGF. Cells were harvested daily, starting at day 2 and continuing until day 8, for reverse transcription-polymerase chain reaction and Western blot. There was no evidence of expression of c-mpl and VEGF receptor on the gene or protein level until day 8, when the formation of well-established hematopoietic colonies began. This correlated with the formation of CD34+/CD31- negative progenitors, mostly found in blast-forming units-erythroid-like colonies. We concluded that TPO and VEGF play an important synergistic role in the formation of early ES-derived hematopoietic progenitors that occurs through the c-mpl and VEGF receptors. Disclosure of potential conflicts of interest is found at the end of this article.

Differentiating embryonic stem (ES) cells are increasingly emerging as an important source of hematopoietic progenitors with a potential to be useful for both basic and clinical research applications. It has been suggested that dexamethasone facilitates differentiation of ES cells towards erythrocytes but the mechanism responsible for sequential expression of genes regulating this process are not well-understood. Therefore, we in vitro induced differentiation of murine ES cells towards erythropoiesis and studied the sequential expression of a set of genes during the process. We hypothesized that dexamethasone-activates its cognate nuclear receptors inducing up-regulation of erythropoietic genes such as GATA-1, Flk-1, Epo-R, and direct ES cells towards erythropoietic differentiation. ES cells were cultured in primary hematopoietic differentiation media containing methyl-cellulose, IMDM, IL-3, IL-6, and SCF to promote embryoid body (EB) formation. Total RNA of day 3, 5, and 9-old EBs was isolated for gene expression studies using RT-PCR. Cells from day 9 EBs were subjected to secondary differentiation using three different cytokines and growth factors combinations: (1) SCF, EPO, dexamethasone, and IGF; (2) SCF, IL-3, IL-6, and TPO; and, (3) SCF IL-3, IL-6, TPO, and EPO. Total RNA from day 12 of secondary differentiated ES cells was isolated to study the gene expression pattern during this process. Our results demonstrate an up-regulation of GATA-1, Flk-1, HoxB-4, Epo-R, and globin genes (alpha-globin, betaH-1 globin, beta-major globin, epsilon -globin, and zeta-globin) in the 9-day-old EBs, whereas, RNA from 5-day-old EBs showed expression of HoxB-4, epsilon-globin, gamma-globin, betaH1-globin, and Flk-1. Three-day-old EBs showed only HoxB-4 and Flk-1 gene expression and lacked expression of all globin genes. These findings indicate that erythropoiesis-specific genes are activated later in the course of differentiation. Gene expression studies on the ES cells of secondary EB origin cultured in media containing dexamethasone showed a down-regulation of GATA-3 and an up-regulation of GATA-1, Flk-1, and Epo-R in comparison to the two other cytokines and growth factor combinations containing media. The secondary differentiation also showed an enhanced production of erythrocytic precursors in dexamethasone containing media in comparison to that in the control media. Our results indicate that dexamethasone can prove to be an effective agent which can be employed to enhance differentiation towards erythrocytic progenitors from ES cells.

An understanding of feasibility of implanting embryonic stem cells (ESCs), their behavior of migration in response to lesions induced in brain tissues, and the mechanism of their in vivo differentiation into neighboring neural cells is essential for developing and refining ESC transplantation strategies for repairing damages in the nervous system, as well as for understanding the molecular mechanism underlying neurogenesis. We hypothesized that damaged neural tissues offer a niche to which injected ESCs can migrate and differentiate into the neural cells. We inflicted damage in the murine (C57BL/6) brain by injecting phosphate-buffered saline into the left frontal and right caudal regions and confirmed neural damage by histochemistry. Enhanced yellow fluorescent protein-expressing ESCs were injected into the nondamaged left caudal portion of the brain. Using immunohistochemistry and fluorescent microscopy, we observed migration of ESCs from the injection site (left caudal) to the damaged site (right caudal and left frontal). Survival of the injected ESCs was confirmed by the real-time polymerase chain reaction analysis of stemness genes such as Oct4, Sox2, and FGF4. The portions of the damaged neural tissues containing ESCs demonstrated a fourfold increase in expression of these genes after 1 week of injection in comparison with the noninjected ESC murine brain, suggesting proliferation. An increased level of platelet-derived growth factor receptor demonstrated that ESCs responded to damaged neural tissues, migrated to the damaged site of the brain, and proliferated. These results demonstrate that undifferentiated ESCs migrate to the damaged regions of brain tissue, engraft, and proliferate. Thus, damaged brain tissue provides a niche that attracts ESCs to migrate and proliferate.

We assessed remission rates and toxicity in 24 consecutive elderly (age>or=60) patients with untreated Acute myeloid leukemia (AML) who received the anthracycline-free combination of fludarabine, cytosine arabinoside and G-CSF (FLAG) as initial induction chemotherapy at our center. CR was achieved following one cycle of FLAG in 14 patients (58%). Another four patients cleared blasts from their bone marrow by day 30 without complete platelet recovery. Three patients died from infections prior to neutrophil recovery (12%). No other grade 3/4 toxicities and no clinically significant mucositis were seen. No significant association was found between age, WBC and cytogenetic risk group with likelihood of achieving CR. Fifteen patients proceeded to consolidation therapy and seven patients received a stem cell transplant (six autologous, one allogeneic). Primary induction with FLAG in elderly AML patients achieves a high remission rate without prohibitive mucosal or cardiac toxicity and may thus be considered as an alternative to standard anthracycline-based regimens in this setting.

Two topics are presented in this review. In the first section, we review data regarding the effects of the endocannabinoids (eCBs) and cannabinoid receptors on neuroimmune function. The function of eCBs in the interaction between the immune system and the central nervous system (CNS) is of particular interest, since the CNS itself is a rich source of eCBs while being exquisitely sensitive to inflammation. There are several sites at which cannabinoids can influence neuroinflammation. Microglial cells express both CB receptors and make eCBs. Activation of CB receptors on these cells seems to promote migration and proliferation but to reduce activation to macrophages. In several neurodegenerative diseases, up-regulation of microglial CB2 receptors have been observed. It is our hypothesis that microglial CB receptor activity is anti-inflammatory and could be exploited to manipulate neuroinflammatory processes with a minimum of unwanted effects. The second topic discussed suggests that the eCB/CB1 receptor pair is involved in the responses of animals to acute, repeated and variable stress. The roles of this pair are complex and dependent upon previous stress, among other things. Dysfunctional responding to stress is a component of several human neuropsychiatric disorders, including anxiety and panic disorders, post-traumatic stress disorders, premenstrual dysphoria and quite possibly, drug abuse. While it is too early to say with certainty, it is very possible that either inhibition or potentiation of endocannabinoid signaling will be an efficacious novel therapeutic approach to more than one human psychiatric disease.

The function and plasticity of the developing immune system during embryonic life has been central to immunological thinking for half a century. A classical view is that antigen encountered during fetal life induces a state of acquired immunological tolerance. However, the ability to develop T cell immune responses during the perinatal period would be of great importance against intracellular pathogens. Recent experiments have challenged this notion and shown that neonatal tolerance can be circumvented by extrinsic immunological manipulations. Here, we used DNA immunization targeted at B lymphocytes to induce a CD4 T cell response that could be measured 2 weeks after birth. We conclude that T cell immunity can be programmed in utero by manipulating the parameters of the immune response in the fetal environment. Furthermore, our data suggest that under appropriate conditions the fetal immune system can be programmed to immunity.

One of the most important issues in stem cell research is to understand the regulatory mechanisms responsible for their differentiation. An extensive understanding of mechanism underlying the process of differentiation is crucial in order to prompt stem cells to perform a particular function after differentiation. To elucidate the molecular mechanisms responsible for the hematopoietic differentiation of embryonic stem cells (ESCs), we investigated murine ES cells for the presence of hematopoietic lineage markers as well as Wnt signaling pathway during treatments with different cytokines alone or in combination with another. Here we report that Wnt/beta-catenin signaling is down-regulated in hematopoietic differentiation of murine ES cells. We also found that differentiation induced by the interleukin-3, interleukin-6, and erythropoietin combinations resulted in high expression of CD3e, CD11b, CD45R/B220, Ly-6G, and TER-119 in differentiated ES cells. A high expression of beta-catenin was observed in two undifferentiated ES cell lines. Gene and protein expression analysis revealed that the members downstream of Wnt in this signaling pathway including beta-catenin, GSK-3beta, Axin, and TCF4 were significantly down-regulated as ES cells differentiated into hematopoietic progenitors. Our results show that the Wnt/beta-catenin signaling pathway plays a role in the hematopoietic differentiation of murine ESCs and also may support beta-catenin as a crucial factor in the maintenance of ES cells in their undifferentiated state.

Background: Clinical application of in utero transplantation (IUT) in human fetuses with intact immune systems resulted in a very low level of donor chimerism. In this study, we examined whether the fetal immune system early in the second trimester of pregnancy (13.5 dpc) can initiate immune tolerance for major histocompatibility complex (MHC)-mismatched embryonic stem (ES) cells. We also examined whether immune tolerance mechanisms respond differently to ontogenetically different stem cells., Methods: MHC-mismatched ES, fetal liver (FL), and bone-marrow (BM) cells (H-2kd) at 1 x 10(9) cells/kg fetal body weight were injected intraperitoneally into 13.5 dpc BALB/c fetuses (H-2Kd). Peripheral chimerism was determined in blood by flow cytometry (sensitivity< or =0.1%) at monthly intervals. Donor-specific immune responses were determined by cytotoxic lymphocyte (CTL) assay, mixed lymphocyte reaction, and T helper (Th)1 and Th2 cytokine assays. Chimeric mice at the age of 9 months received postnatal boosts (PB) with minimal conditioning of 200 cGy by intravenous injection of 1 x 10(9) of the corresponding cells/kg body weight., Results: After IUT with ES, FL, or BM cells, the level of peripheral chimerism within the first 9 months of life was 0% to 0.4%. PB with 1 x 10(9)/kg of corresponding cells resulted in a decrease in the peripheral chimerism to 0% within 2 weeks of PB. CTL and cytokine assays before and after PB demonstrated a shift toward immunity., Conclusions: Immunologic tolerance was not achieved after IUT of murine fetuses at 13.5 dpc with MHC-mismatched ES cells, and only a low level chimerism was achieved.

Between January 1996 and July 2002, 72 patients with non-Hodgkin's lymphoma or Hodgkin's disease underwent high-dose chemotherapy with autologous stem cell transplant conditioned with either cyclophosphamide, etoposide, carmustine (CEB) or carmustine, etoposide, cytarabine, melphalan (BEAM) at a single institution. In all, 52 patients received CEB and 20 patients received the BEAM regimen. Patient characteristics that were significantly different between the two groups are tumor grade and extranodal involvement (P = 0.0196, 0.0341, respectively). Regimen-related toxicities examined yielded only diarrhea occurring at a higher rate in the BEAM group (81 vs 51%, P = 0.0026), although cases were milder (92 vs 57%). Patients treated with CEB developed mucositis at a slightly higher rate (79%) than patients treated with BEAM (75%), but this difference did not reach statistical significance. However, the mucositis that occurred within the BEAM group was predominately mild (67%) in contrast to the predominance of moderate to severe cases in the CEB group (74%). In addition, patients treated with CEB required growth factor support for a longer time than patients treated with BEAM (P = 0.0399). Response rates were high in both groups, with trends favoring the BEAM group. Overall survival was higher after treatment with BEAM than with CEB (84 vs 60%).

The phage is used as a scaffold to display recombinant libraries of peptides, which provides the means to rescue and amplify peptides that bind target macromolecules. Many reports showed that the T7 phage display method can be used to obtain a ligand-binding peptidefor tissue-targeted therapies in adult animals. In utero tissue targeting of fetal tissues may help in the correction of many genetic and metabolic diseases. Here we demonstrate the distribution and detection of T7 phage displaying the C-X7-C peptide library in mouse fetal tissues after systemic injection of T7 phage into pregnant mouse tail vein. T7 phage was recovered from fetal tissues 15 min after injection of T7 phage. Our results suggest that T7 phage may be a useful tool in selecting the tissue-specific ligand-binding peptide for fetal tissues. This approach may be helpful in designing in utero tissue-targeted therapies.

Curcumin, a phenolic compound from the rhizome of the plant Curcuma longa has anti-inflammatory, antioxidant and anti-cancer activities. Although the precise mode of action of this compound is not yet elucidated, studies have shown that chemo-preventive action of curcumin might be due to its ability to induce apoptosis and to arrest cell cycle. This study investigated the cellular and molecular changes induced by curcumin leading to the induction of apoptosis in human lung cancer cell lines-A549 and H1299. A549 is p53 proficient and H1299 is p53 null mutant. The lung cancer cells were treated with curcumin (0-160 microM) for 12-72 h. Curcumin inhibited the growth of both the cell lines in a concentration dependent manner. Growth inhibition of H1299 cell lines was both time and concentration dependent. Curcumin induced apoptosis in both the lung cancer cell lines. A decrease in expression of p53, bcl-2, and bcl-X(L) was observed after 12 h exposure of 40 microM curcumin. Bak and Caspase genes remained unchanged up to 60 microM curcumin but showed decrease in expression levels at 80-160 microM. The data also suggest a p53 independent induction of apoptosis in lung cancer cells.

Cytosolic alanine aminotransferase (c-AAT) was purified up to 203- and 120-fold, from the liver of two freshwater teleosts Clarias batrachus (air-breathing, carnivorous) and Labeo rohita (water-breathing, herbivorous), respectively. The enzyme from both fish showed similar elution profiles on a DEAE-Sephacel ion exchange column. SDS-PAGE of purified enzymes revealed two subunits of 54 and 56 kDa, in both fish. The apparent Km values for l-alanine were 18.5+/-0.48 and 23.55+/-0.60 mM, whereas for 2-oxoglutarate the Km values were observed to be 0.29+/-0.023 and 0.33+/-0.028 mM for the enzyme from C. batrachus and L. rohita, respectively. With l-alanine as substrate, aminooxyacetic acid was found to act as a competitive inhibitor with KI values of 6.4 x 10(-4) and 3.4 x 10(-4) mM with c-AAT of C. batrachus and L. rohita, respectively. However, when 2-oxoglutarate was used as substrate, aminooxyacetic acid showed uncompetitive inhibition with similar KI values for purified c-AAT from both fish. Temperature and pH profiles of the enzyme did not show any marked differences between the two fish examined. These results suggest that liver c-AAT, isolated from these two fish species adapted to different modes of life, remain unaltered structurally. However, at the kinetic level, liver c-AAT from C. batrachus exhibits significantly higher affinity for the substrate l-alanine and decreased affinity for its metabolic inhibitor, in comparison to that of the enzyme purified from L. rohita. Such functional changes seem to be of physiological significance and also provide preliminary evidence for subtle changes in the enzyme as a mark of metabolic adaptation in the fish to different physiological demands.

Phage display is a powerful method to study organ and tissue specific addresses. As part of our studies on the in vivo panning of tissue-homing peptide libraries, we examined the survival of T7 phage in the blood of C57BL/6J mice to estimate the half-life of T7 phage and the factors responsible for its inactivation. Amplified and purified T7 phage particles with or without random peptide library inserts were injected intravenously into the tail vein of wild-type (C57BL/6J) and immunocompromized (C57BL/6J) female mice. In wild-type mice, both the parent phage as well as phage carrying a peptide library were eliminated quickly from the blood, with only approximately 1% survival of detectable infectious phage after 60 min of injection. In SCID (C57BL/6J-Prkdc(scid)) mice, phage titers were stable over the same period of time with or without peptide library, suggesting a role for either B- or T cells or both in phage inactivation. The presumed role of B cell was indicated by demonstration of stable phage in the B-cell deficient mouse (C57BL/10-Igh-6(tm1Cgn)). In other immunocompromized mice, the phage titers were unstable, similar to that found in wild-type mice. In no case, was there a difference between phage with or without random peptide library. These data indicate that the presence of random C-X7-C peptides on the T7 phage coat protein does not affect the clearance of the phage in murine blood. Most likely, host immune factors play a role in the neutralization of T7 phage in blood by reacting with B-cell dependent immunoglobin.

Background: The adequacy of HPC collection for BMT is typically assessed by the number of CD34 cells. However, during a series of leukapheresis procedures (LP) the CD34 value on the final HPC product may not be available for testing until late evening, sometimes resulting in additional, retrospectively unnecessary, LP in order to ensure an adequate HPC collection (>5x10(6) CD34/kg). We hypothesized that an estimate of the CD34 content of HPC products prior to 16:00 h on the day of LP would permit improved HPC collection planning. We therefore assessed the effectiveness of predicting the total amount of CD34 cells that would be collected in a given LP by either (a) the concentration of CD34 cells/microL in peripheral blood prior to LP (pre-CD34) or (b) the predicted total amount of CD34 cells to be collected based on sampling the LP product at the mid-point of each LP. We also compared the number of LP per patient and total HPC collected for the study group with data from the previous calendar year., Methods: Allogeneic and autologous BMT donors who completed a 20-L HPC collection between September 2002 and February 2003 were eligible. CD34 cells were measured on blood drawn prior to LP and from the HPC product at the mid-point (10 L) of LP. The CD34 content of the final LP was predicted by doubling the value of total CD34 cells at the mid-run (MRp-CD34). The MRp-CD34/kg and the cumulative CD34/kg collected were made available before 16:00 h and used to determine the need for additional LP. The true CD34 content of each HPC collection was also measured from the final product the next day (CD34-FP)., Results: A 20-L LP was completed and data were available from 31 patients and nine allogeneic donors who underwent a total of 85 LP for diagnoses, including 11 myeloma, 10 lymphoma, seven HD, three acute leukemia and five others. The mean (range) and correlation (R2) vs. the CD34-FP were, for pre-CD34, 54 CD34/microL (0.3-232), R2=0.66 (P<0.01), and for MRp-CD34, 3.2x10(6) CD34/kg (0.04-22.48), R2=0.90 (P<0.01). The mean number of CD34/kg collected per LP in the patients/donors was 3.4x10(6) CD34/kg (0.05-18.94). The median number of CD34 cells employed for transplant in the study group vs. controls (5.7 vs. 5.6x10(6)/kg) and the time to engraftment of neutrophils (12 vs. 11 days) and platelets (12 vs. 12 days) was similar to historical controls. However, the study group had a significantly lower median number of LP (three vs. two; P<0.02) to obtain the required collection of 5x10(6) CD34 cells/kg., Discussion: Both the pre-CD34 and the MRp-CD34 were significantly correlated with CD34-FP. However, the CD34-FP was more reliably predicted by MRp-CD34. Early availability of mid-run CD34 values was associated with a significant reduction in the number of LP required to collect 5x10(6) CD34 cells/kg, without reduction in the number of CD34 cells for transplant or prolongation of days to neutrophil or platelet engraftment.

A 38-year-old woman with agnogenic myeloid metaplasia complicated by the poor prognostic factors of severe osteosclerosis, prominent hepatosplenomegaly, and profound anemia was treated with FLAG chemotherapy to decrease her organomegaly before undergoing a nonmyeloablative allogeneic stem cell transplant from a matched-sibling donor. The patient's pre- and post transplant course were complicated by an autoimmune disorder and her post transplant course was complicated by severe hepatic and gastrointestinal GVHD. A technetium-99m sulfur colloid scan 4 months post transplant and bone marrow studies 8 months post transplant demonstrated intramedullary hematopoiesis, complete resolution of marrow fibrosis, and partial resolution of osteosclerosis.

At Northwestern University, a phase I/II trial of hematopoietic stem cell transplant (HSCT) for systemic lupus erythematosus (SLE) has shown promising results. A phase III HSCT trial is being developed to confirm efficacy of HSCT vs continuing the currently accepted standard of care, intravenous pulse cyclophosphamide.