Evidence of both ontogeny and transplant dose-regulated expansion of hematopoietic stem cells in vivo

Recent assessment of the long-term repopulating activity of defined subsets of hematopoietic cells has offered new insights into the characteristics of the transplantable stem cells of this system; however, as yet, there is very little known about mechanisms that regulate their self-renewal in vivo. We have now exploited the ability to quantitate these cells using the competitive repopulating unit (CRU) assay to identify the role of both intrinsic (ontological) and extrinsic (transplanted dose-related) variables that may contribute to the regulation of CRU recovery in vivo. Ly5.1 donor cells derived from day-14.5 fetal liver (FL) or the bone marrow (BM) of adult mice injected 4 days previously with 5-fluorouracil were transplanted at doses estimated to contain 10, 100, or 1,000 long-term CRU into irradiated congenic Ly5.2 adult recipient mice. Eight to 12 months after transplantation, there was a complete recovery of BM cellularity and in vitro clonogenic progenitor numbers and a nearly full recovery of day-12 colony-forming unit-spleen numbers irrespective of the number or origin of cells initially transplanted. In contrast, regeneration of Ly5.1+ donor-derived CRU was incomplete in all cases and was dependent on both the origin and dose of the transplant, with FL being markedly superior to that of adult BM. As a result, the final recovery of the adult marrow CRU compartment ranged from 15% to 62% and from 1% to 18% of the normal value in recipients of FL and adult BM transplantation, respectively, with an accompanying maximum CRU amplification of 150- fold for recipients of FL cells and 15-fold for recipients of adult BM cells. Interestingly, the extent of CRU expansion from either source was inversely related to the number of CRU transplanted. These data suggest that recovery of mature blood cell production in vivo may activate negative feedback regulatory mechanisms to prematurely limit stem cell self-renewal ability. Proviral integration analysis of mice receiving retrovirally transduced BM cells confirmed regeneration of totipotent lymphomyeloid repopulating cells and provided evidence for a greater than 300-fold clonal amplification of a single transduced stem cell. These results highlight the differential regenerative capacities of CRU from fetal and adult sources that likely reflect intrinsic, genetically defined determinants of CRU expansion but whose contribution to the magnitude of stem cell amplification ultimately obtained in vivo is also strongly influenced by the initial number of CRU transplanted. Such findings set the stage for attempts to enhance CRU regeneration by administration of agents that may enable full expression of regenerative potential or through the expression of intracellular gene products that may alter intrinsic regenerative capacity.     

Detection of platelet-directed immunoglobin G in sera using the peroxidase-anti-peroxidase (PAP) slide technique

An immunohistochemical procedure for the detection of immunoglobulin G adherent to platelets is described. The peroxidase anti-peroxidase method is used to detect antibody activity directed against platelets from normal donors in the sera from 305 individuals. These subjects were divided into three groups: group 1, patients referred for tissue typing; group 2, healthy normal females; group 3, healthy normal males. In group 1, 28% of the sera were found to be positive; in most of these a history of prior transfusions was obtained. In group 2, 7.4% were found to be positive, most having previous pregnancies. Only 1% were found to be positive in group 3, and no reason for presensitization was found. Results from the indirect immunofluorescence technique served as a control and as a means to compare the sensitivity. Under the conditions chosen, the peroxidase anti-peroxidase test was two to eight times more sensitive than the immunofluorescence technique. Specificity of the peroxidase anti-peroxidase technique was demonstrated using a monospecific anti-PLA1 antiserum. It is concluded that the peroxidase anti-peroxidase slide technique may be a useful tool in the study of platelet-related immunophenomena.     

Demonstration of a blood-bone marrow barrier to macrophage colony- stimulating factor

Several previous studies suggested that murine macrophage colony- stimulating factor (CSF-1) might have impaired access to hematopoietic cells in the marrow. The apparent lack of hematopoietic responses to exogenous CSF and the finding of available or unoccupied CSF receptors despite saturating CSF levels in the serum led to studies of a potential blood-bone marrow barrier for this factor. Groups of mice were injected with pure unlabeled CSF-1 by either intravenous (IV) or intraperitoneal (IP) routes. Marrow and spleen cells were obtained at intervals after injection, held at 0 degree C, and assessed for changes in binding of 125I-CSF. Saturation of all available CSF receptors is achieved in vitro with 100 to 150 U CSF/mL. Despite achieving serum levels of 5,000 to 7,000 U/mL after IV injection of 25,000 units of CSF, less than 50% of the marrow receptors and less than 85% of the splenic receptors were saturated or downregulated. The decline in receptor availability was transient, with return of receptor sites in two to four hours. Increasing the IV dose to 125,000 units increased serum CSF values to approximately 20,000 U/mL and led to a virtual disappearance of available receptors for two to three hours. When administered IP, only approximately 40% of marrow and 80% of splenic receptors were affected for two hours. It was necessary to increase the dose of CSF to 250,000 units IP to saturate or downregulate receptors for three to four hours after injection. These observations indicate a marked blood-bone marrow barrier and lesser blood-spleen barrier for the transfer of serum CSF to responsive hematopoietic cells in vivo.     

Fatigue Intervention by Nurses Evaluation – The FINE Trial. A randomised controlled trial of nurse led self-help treatment for patients in primary care with chronic fatigue syndrome: study protocol. [ISRCTN74156610]

Background

A major change has occurred in the last few years in the therapeutic approach to patients presenting with all forms of acute coronary syndromes. Whether or not these patients present initially to tertiary cardiac care centers, they are now routinely referred for early coronary angiography and increasingly undergo percutaneous revascularization. This practice is driven primarily by the angiographic image and technical feasibility. Concomitantly, there has been a decline in expectant or ischemia-guided medical management based on specific clinical presentation, response to initial treatment, and results of noninvasive stratification. This 'tertiarization' of acute coronary care has been fuelled by the increasing sophistication of the cardiac armamentarium, the peer-reviewed publication of clinical studies purporting to show the superiority of invasive cardiac interventions, and predominantly supporting (non-peer-reviewed) editorials, newsletters, and opinion pieces.

Discussion

This review presents another perspective, based on a critical reexamination of the evidence. The topics addressed are: reperfusion treatment of ST-elevation myocardial infarction; the indications for invasive intervention following thrombolysis; the role of invasive management in non-ST-elevation myocardial infarction and unstable angina; and cost-effectiveness and real world considerations. A few cases encountered in recent practice in community and tertiary hospitals are presented for illustrative purposes The numerous and far-reaching scientific, economic, and philosophical implications that are a consequence of this marked change in clinical practice as well as healthcare, decisional and conflict of interest issues are explored.

Summary

The weight of evidence does not support the contemporary unfocused broad use of invasive interventional procedures across the spectrum of acute coronary clinical presentations. Excessive and unselective recourse to these procedures has deleterious implications for the organization of cardiac health care and undesirable economic, scientific and intellectual consequences. It is suggested that there is need for a new equilibrium based on more refined clinical risk stratification in the treatment of patients who present with acute coronary syndromes.     

Cytomegalovirus-induced necrotizing and crescentic glomerulonephritis in a renal transplant patient

A 35-year-old black man with end-stage renal disease from biopsy-proven focal segmental glomerulosclerosis developed worsening function of his renal allograft 160 days after living related donor renal transplantation. Renal biopsy showed necrotizing and crescentic glomerulonephritis (NCGN) and presence of intraglomerular viral inclusions confirmed by immunocytochemical stain and in situ hybridization techniques to be cytomegaloviral in origin. Electron microscopy showed no immune complexes, and workup for other causes of NCGN was negative. The patient was treated with ganciclovir without other changes in his immunosuppressive regimen. After 8 weeks of ganciclovir therapy, a second renal transplant biopsy showed resolution of the glomerular process and disappearance of the cytomegalovirus (CMV) inclusions. The resolution of the glomerular process with treatment for CMV infection, and without other change in therapy, strongly supports a causative link between CMV and NCGN in this patient. This case represents the first report of CMV-associated NCGN in a renal transplant patient.