Kinetics of peripheral blood mononuclear cell mobilization with chemotherapy and/or granulocyte-colony-stimulating factor: implications for yield of hematopoietic progenitor cell collections

BACKGROUND: Peripheral blood progenitor cells (PBPCs) are commonly collected and used to reconstitute hematopoiesis after high-dose chemotherapy. However, strategies for optimal collection and assessment of leukapheresis components are not standardized. STUDY DESIGN and METHODS: Hematopoietic progenitor cell assays were performed on 369 leukapheresis components collected from 95 patients who had received doxorubicin-based chemotherapy and/or granulocyte-colony-stimulating factor (G-CSF). Precollection patient hematologic values, leukapheresis collection values, component hematopoietic progenitor cell assays, and patient outcome measures were summarized. The kinetics of mononuclear cell (MNC) and PBPC mobilization were assessed among four patient groups. RESULTS: Patient group was a significant predictor of the peripheral blood MNC count on the day of collection (p<0.0001), and that value was a significant predictor of granulocyte-macrophage– colony-forming unit (CFU-GM) yield (p<0.0001). This relationship between the peripheral blood MNC count on the day of collection and CFU- GM yield differed according to patient group (p<0.0001). CFU-GM made up a larger fraction of peripheral blood MNCs collected from patients who received chemotherapy plus G-CSF than collected from those who received G-CSF alone. Moreover, the peripheral blood MNC count and the corresponding CFU-GM yield increased significantly on consecutive days of collection in patient groups receiving chemotherapy and G-CSF but were unchanged or decreased in patients receiving G-CSF alone. CONCLUSION: The relationship between peripheral blood MNC count and leukapheresis component CFU-GM yield differed significantly between patients who received chemotherapy and G-CSF and those who received G- CSF alone for the mobilization of PBPCs. Patient peripheral blood MNC count and component CFU-GM yield are useful for both assessing and suggesting revisions to PBPC mobilization and collection strategies.     

Human MSH2 binds to trinucleotide repeat DNA structures associated with neurodegenerative diseases

The expansion of trinucleotide repeat sequences is associated with several neurodegenerative diseases. The mechanism of this expansion is unknown but may involve slipped-strand structures where adjacent rather than perfect complementary sequences of a trinucleotide repeat become paired. Here, we have studied the interaction of the human mismatch repair protein MSH2 with slipped-strand structures formed from a triplet repeat sequence in order to address the possible role of MSH2 in trinucleotide expansion. Genomic clones of the myotonic dystrophy locus containing disease-relevant lengths of (CTG)n x (CAG)n triplet repeats were examined. We have constructed two types of slipped-strand structures by annealing complementary strands of DNA containing: (i) equal numbers of trinucleotide repeats (homoduplex slipped structures or S-DNA) or (ii) different numbers of repeats (heteroduplex slipped intermediates or SI-DNA). SI-DNAs having an excess of either CTG or CAG repeats were structurally distinct and could be separated electrophoretically and studied individually. Using a band-shift assay, the MSH2 was shown to bind to both S-DNA and SI-DNA in a structure- specific manner. The affinity of MSH2 increased with the length of the repeat sequence. Furthermore, MSH2 bound preferentially to looped-out CAG repeat sequences, implicating a strand asymmetry in MSH2 recognition. Our results are consistent with the idea that MSH2 may participate in trinucleotide repeat expansion via its role in repair and/or recombination.      

Catecholamine synthesis and metabolism in the central nervous system of mice lacking α2-adrenoceptor subtypes

Background and purpose:

This study investigates the role of α₂-adrenoceptor subtypes, α_2A, α_2B and α_2C, on catecholamine synthesis and catabolism in the central nervous system of mice.

Experimental approach:

Activities of the main catecholamine synthetic and catabolic enzymes were determined in whole brains obtained from α_2A-, α_2B- and α_2C-adrenoceptor knockout (KO) and C56Bl\7 wild-type (WT) mice.

Key results:

Although no significant differences were found in tyrosine hydroxylase activity and expression, brain tissue levels of 3,4-dihydroxyphenylalanine were threefold higher in α_2A- and α_2C-adrenoceptor KO mice. Brain tissue levels of dopamine and noradrenaline were significantly higher in α_2A and α_2CKOs compared with WT [WT: 2.8 ± 0.5, 1.1 ± 0.1; α_2AKO: 6.9 ± 0.7, 1.9 ± 0.1; α_2BKO: 2.3 ± 0.2, 1.0 ± 0.1; α_2CKO: 4.6 ± 0.8, 1.5 ± 0.2 nmol·(g tissue)⁻¹, for dopamine and noradrenaline respectively]. Aromatic L-amino acid decarboxylase activity was significantly higher in α_2A and α_2CKO [WT: 40 ± 1; α_2A: 77 ± 2; α_2B: 40 ± 1; α_2C: 50 ± 1, maximum velocity (V_max) in nmol·(mg protein)⁻¹·h⁻¹], but no significant differences were found in dopamine β-hydroxylase. Of the catabolic enzymes, catechol-O-methyltransferase enzyme activity was significantly higher in all three α₂KO mice [WT: 2.0 ± 0.0; α_2A: 2.4 ± 0.1; α_2B: 2.2 ± 0.0; α_2C: 2.2 ± 0.0 nmol·(mg protein)⁻¹·h⁻¹], but no significant differences were found in monoamine oxidase activity between all α₂KOs and WT mice.

Conclusions and implications:

In mouse brain, deletion of α_2A- or α_2C-adrenoceptors increased cerebral aromatic L-amino acid decarboxylase activity and catecholamine tissue levels. Deletion of any α₂-adrenoceptor subtypes resulted in increased activity of catechol-O-methyltransferase. Higher 3,4-dihydroxyphenylalanine tissue levels in α_2A and α_2CKO mice could be explained by increased 3,4-dihydroxyphenylalanine transport.     

MR fluoroscopy: initial clinical studies

Magnetic resonance (MR) fluoroscopy is a method for high-speed MR image acquisition with the goals of short acquisition time per image (500 msec or less), high image rate (10 images or more per second), and high-speed image reconstruction (150 msec or less from data acquisition to image display). The authors present their results with the first two goals in volunteers. MR fluoroscopic image data were acquired with a limited flip angle pulse sequence with reduced repetition times (TRs) and fewer phase encodings used per image. The sequence was applied continuously, and images were formed by updating one set of data with data from the most recently taken measurements. Sample head images were generated with TR/echo times as small as 11/5.5 msec and 48 phase encodings for a total acquisition time of about 500 msec. Images were acquired while the volunteer flexed his head. Artifacts from the motion became less evident on images as progressively shorter acquisition times were used.     

Tetralogy of Fallot: MR findings

Surgical treatment of patients with tetralogy of Fallot requires accurate definition of all anatomic structures, particularly the central pulmonary arteries. Magnetic resonance (MR) images of 22 patients with tetralogy of Fallot were studied to assess their usefulness in providing information regarding the spectrum of anatomic abnormalities in this condition. MR findings were compared with information obtained at catheterization (in 16 patients) and at surgery (in nine patients), both of which were performed within 3 months of MR imaging. Ventricular chamber enlargement and wall hypertrophy were clearly delineated in most of the 17 patients who were examined before definitive surgical repair, and ventricular septal defects were visualized in all 17. Palliative systemic-to-pulmonary shunts were visualized in 11 patients and could be evaluated for patency. Most important, the morphology and size of the right ventricular outflow tract and central pulmonary arteries could be accurately assessed. Pulmonary artery measurements obtained from MR images demonstrated excellent correlation with angiographic measurements. In six patients examined after complete surgical repair, MR images accurately reflected changes in pulmonary artery outflow tract morphology and complications, such as residual pulmonary artery stenosis and thrombosis. The findings suggest that MR imaging can complement or obviate catheterization in the evaluation of tetralogy of Fallot with regard to suitability for definitive surgical repair.     

Hematopoietic and fatty bone marrow distribution in the normal and ischemic hip: new observations with 1.5-T MR imaging

The conversion of hematopoietic to fatty marrow is known to correlate with physiologic decreases in intramedullary blood flow. To determine whether the chronology of conversion is altered in patients with hip ischemia, T1-weighted magnetic resonance (MR) images of the hips in 50 healthy people and 27 with documented avascular necrosis (AVN) were reviewed. The distribution of fatty (high-signal) versus hematopoietic (low-signal) marrow was noted with respect to age. All patients had fatty marrow in the femoral capital epiphysis and greater trochanter. Hematopoietic intertrochanteric marrow was seen in 95% (80 of 84) of femurs in control subjects less than 50 years old, but in only 12.5% (two of 16) of those in control subjects older than 50 years (P less than .005). Only 33% (19 of 57) of patients less than 50 years with AVN had predominantly hematopoietic intertrochanteric marrow (P less than .005). The early conversion to fatty marrow in most patients with AVN as depicted by MR imaging may be an effect of decreased vascularity of the proximal femur and may allow the identification of patients at increased risk for AVN.     

Detection of DNA adducts of acetaldehyde in peripheral white blood cells of alcohol abusers

The effect of alcohol drinking on the formation of DNA adducts of acetaldehyde, the primary oxidative metabolite of ethanol, was investigated in humans. DNA was isolated from granulocytes and lymphocytes from 24 alcoholic patients and 12 control subjects. DNA adduct levels were measured by 32P-postlabelling using reversed-phase HPLC with on-line detection of radioactivity. A large interindividual variation in adduct levels was observed. The average adduct levels in granulocyte and lymphocyte DNA from alcoholic patients were 3.4 +/- 3.8 and 2.1 +/- 0.8 adducts/10(7) nucleotides (n = 24), respectively. These levels were 13- and 7-fold higher than the corresponding levels in control subjects (P<0.001). The average adduct level in granulocyte DNA from alcoholic patients was 60% higher than in lymphocyte DNA (P<0.01). Our results, in conjunction with the genotoxicity of acetaldehyde, thus suggest the formation of DNA adducts of acetaldehyde as a plausible mechanism explaining the involvement of alcohol drinking in carcinogenesis.