Human erythrocyte protein 4.2 deficiency associated with hemolytic anemia and a homozygous 40glutamic acid-->lysine substitution in the cytoplasmic domain of band 3 (band 3Montefiore)

Red blood cell (RBC) protein 4.2 deficiency is often associated with a moderate nonimmune hemolytic anemia, splenomegaly, and osmotically fragile RBCs resembling, but not identical to, hereditary spherocytosis (HS). In the Japanese type of protein 4.2 deficiency (protein 4.2Nippon), the anemia is associated with a point mutation in the protein 4.2 cDNA. In this report, we describe a patient with moderate and apparently episodic nonimmune hemolytic anemia with splenomegaly, spherocytosis, osmotically fragile RBCs, reduced whole cell deformability, and abnormally dense cells. Sodium dodecyl sulfate- polyacrylamide gel electrophoresis analysis of the proposita's RBC membrane proteins showed an 88% deficiency of protein 4.2 and a 30% deficiency of glyceraldehyde-3-phosphate dehydrogenase (band 6). Structural and molecular analyses of the proposita's protein 4.2 were normal. In contrast, limited tryptic digestion of the proposita's band 3 showed a homozygous abnormality in the cytoplasmic domain. Analysis of the pedigree disclosed six members who were heterozygotes for the band 3 structural abnormality and one member who was a normal homozygote. Direct sequence analysis of the abnormal band 3 tryptic peptide suggested that the structural abnormality resided at or near residue 40. Sequence analysis of the proposita's band 3 cDNA showed a 232G-->A mutation resulting in a 40glutamic acid-->lysine substitution (band 3Montefiore). Allele-specific oligonucleotide hybridization was used to probe for the mutation in the pedigree, showing that the proposita was homozygous, and the pedigree members who were heterozygous for the band 3 structural abnormality were also heterozygous for the band 3Montefiore mutation. The band 3Montefiore mutation was absent in 26 chromosomes from race-matched controls and in one pedigree member who did not express the band 3 structural abnormality. In coincidence with splenectomy, the proposita's anemia was largely corrected along with the disappearance of most spherocytes and considerable improvements of RBC osmotic fragility, whole cell deformability, and cell density. We conclude that this hereditary hemolytic anemia is associated with the homozygous state for band 3Montefiore (40glutamic acid-->lysine) and a decreased RBC membrane content of protein 4.2. We speculate that band 3 structural abnormalities can result in defective interactions with protein 4.2 and band 6, and in particular, that the region of band 3 containing 40glutamic acid is involved directly or indirectly in interactions with these proteins.     

Fetal liver hematopoietic stem cells as a target for in utero retroviral gene transfer.

Retroviral-mediated gene transfer into hematopoietic precursors often results in only short-term gene transduction in vivo. Loss of the transduced genetic material over time may be caused by the limited ability of retroviral infection to transduce genes into early, pluripotent hematopoietic stem cells. Because fetal liver contains actively proliferating multipotential stem cells that should be more susceptible to retroviral-mediated gene transfer than quiescent cells derived from adult bone marrow, these cells may be an ideal target for gene transduction. Furthermore, physiologic expansion of these cells during development obviates the need for marrow ablation during gene therapy in vivo. We performed in utero gene transfer by injecting high titer replication-defective retrovirus in vivo into the livers of 11, 14, 16, and 18 day gestation rats. After birth, the rats were analyzed for the presence of proviral integration and gene expression. The efficiency of gene transfer into bone marrow cells was greatest in rats infected at day 14 to 16 of gestation. In rats killed at 1 to 26 weeks of age, gene transfer was detected by Southern analysis in 48% and by polymerase chain reaction in 86% of bone marrow samples. The provirus was also detected in white blood cells, the granulocyte-macrophage colony-forming unit, thymus, spleen, liver, and lung. The presence of the transgene in bone marrow and other hematopoietic tissues at 26 weeks of age suggests that early hematopoietic precursors present in the fetal liver are susceptible targets for gene transfer and that these cells become resident in the bone marrow of the adult animal. This model is a new technique for gene transduction into proliferating hematopoietic cells in vivo that avoids bone marrow transplantation and has potential application in the correction of genetic defects in utero.     

Sympathetic nervous system function as measured by I-123 metaiodobenzylguanidine predicts transplant-free survival in heart failure patients with idiopathic dilated cardiomyopathy

BACKGROUND: Heightened activity of the sympathetic nervous system in heart failure patients is a major contributor to disease progression and death. I-123 metaiodobenzylguanidine (MIBG) provides an accurate, noninvasive method to assess cardiac sympathetic nerve activity. METHODS: Thirty-seven patients with New York Heart Association class II, III, or IV heart failure underwent baseline measurement of I-123 MIBG heart-to-mediastinum ratios, maximum oxygen consumption, radionuclide left ventricular ejection fraction, and plasma norepinephrine levels. Patients were followed 48.8+/-8.6 months to endpoints of cardiac death or transplantation. The heart-to-mediastinum ratio of I-123 MIBG activity measured 15 minutes after injection was the only independent predictor of transplant-free survival (P<.0001). I-123 MIBG imaging at 15 minutes identified patients with subsequent cardiac transplantation or death with a sensitivity of 92% and specificity of 72%, whereas the corresponding values for maximum oxygen consumption were 75% and 56%. By Kaplan-Meier survival analysis, the time to a cardiac endpoint was significantly shorter in patients with a 15-minute I-123 MIBG heart-to-mediastinum ratio below the group mean ratio of 1.536, compared with patients with a preserved I-123 MIBG ratio. Maximum oxygen consumption was not predictive of time to cardiac transplant or death. CONCLUSIONS: In this study of patients with congestive heart failure resulting from dilated cardiomyopathy, a 15-minute heart-to-mediastinum ratio of I-123 MIBG activity provided more accurate prediction of cardiac transplantation or death than other standard clinical tests.     

A prospective study of oesophageal 24-h ambulatory pH monitoring in patients with functional dyspepsia

BACKGROUND: The diagnostic utility of 24-h oesophageal ambulatory pH monitoring in patients with functional dyspepsia has not been well established. AIMS: We performed a prospective study of oesophageal pH monitoring in patients with functional dyspepsia in order to assess whether a positive pH test might predict response to proton pump inhibitor therapy in a subset of functional dyspepsia patients. PATIENTS: Forty Helicobacter pylori-negative functional dyspepsia patients (35 males and 5 females, mean age (+/-S.E.M.) of 54+/-2.4 years) with predominantly unspecified dyspepsia subtype and normal distal oesophageal biopsies. METHODS: All subjects were randomised in a double-blind fashion to either omeprazole 20 mg/day or placebo daily for four weeks after 24-h pH monitoring. RESULTS: Twenty-four-hour pH monitoring was abnormal in 9 of the 21 patients (43%) in the omeprazole group and 5/19 (26%) of the placebo group (p=NS). Patients who reported symptomatic improvement on the Gastrointestinal Symptom Rating Scale were no more likely to have abnormal scores on pH monitoring than patients who did not have symptomatic response. CONCLUSIONS: Although approximately one-third of functional dyspepsia patients will have abnormal profiles on 24-h ambulatory oesophageal pH monitoring, an abnormal score does not appear to predict response to proton pump inhibitor therapy in patients with unspecified functional dyspepsia.     

Cardiac thrombosis and thromboembolism in chronic chagas' heart disease

A retrospective study of Chagas' heart disease was carried out by a review of 1,345 autopsy reports, with special reference to cardiac thrombus and thromboembolic phenomena. The incidence of cardiac thrombus was higher in cases of heart failure (36%) than in cases of sudden death (15%), higher in heavier hearts, and unrelated to age or sex. The left- and right-sided cardiac chambers were equally affected by thrombus. Endocarditis and blood stasis were considered important factors in the pathogenesis of cardiac thrombus. Thromboembolic phenomena were more common in the systemic circulation but caused relatively more deaths by pulmonary embolism. Fourteen percent of patients with thromboembolic phenomena died from them. Patients with multiple thromboembolic phenomena had a higher risk of death from embolism. Cardiac thrombosis or thromboembolic phenomena, or both, were present in 44% of the cases studied. Prophylactic measures should be taken for these important complications of Chagas' heart disease.     

Primary tumor cells of myeloma patients induce interleukin-6 secretion in long-term bone marrow cultures

Long-term bone marrow cultures (LTBMC) from patients with multiple myeloma (MM) and normal donors were analyzed for immunophenotype and cytokine production. Both LTBMC adherent cells from myeloma and normal donor origin expressed CD10, CD13, the adhesion molecules CD44, CD54, vascular cell adhesion molecule 1, very late antigen 2 (VLA-2), and VLA- 5, and were positive for extracellular matrix components fibronectin, laminin, and collagen types 3 and 4. LTBMC from myeloma patients and normal donors spontaneously secreted interleukin-6 (IL-6). However, levels of IL-6 correlated with the stage of disease; highest levels of IL-6 were found in LTBMC from patients with active myeloma. To identify the origin of IL-6 production, LTBMC from MM patients and normal donors were cocultured with BM-derived myeloma cells and cells from myeloma cell lines. IL-6 was induced by plasma cell lines that adhered to LTBMC such as ARH-77 and RPMI-8226, but not by nonadhering cell lines U266 and FRAVEL. Myeloma cells strongly stimulated IL-6 secretion in cocultures with LTBMC adherent cells from normal donors and myeloma patients. When direct cellular contact between LTBMC and plasma cells was prevented by tissue-culture inserts, no IL-6 production was induced. This implies that intimate cell-cell contact is a prerequisite for IL-6 induction. Binding of purified myeloma cells to LTBMC adherent cells was partly inhibited by monoclonal antibodies against adhesion molecules VLA-4, CD44, and lymphocyte function-associated antigen 1 (LFA-1) present on the plasma cell. Antibodies against VLA-4, CD29, and LFA-1 also inhibited the induced IL-6 secretion in plasma cell-LTBMC cocultures. In situ hybridization studies performed before and after coculture with plasma cells indicated that LTBMC adherent cells produce the IL-6. These results suggest that the high levels of IL-6 found in LTBMC of MM patients with active disease are a reflection of their previous contact with tumor cells in vivo. These results provide a new perspective on tumor growth in MM and emphasize the importance of plasma cell-LTBMC interaction in the pathophysiology of MM.     

Reliability of two pragmatic tools for assessing text neck

Background

There is a hypothesis that the growing use of mobile phones in an inappropriate posture to text and read (text neck) could be a reason for the increasing prevalence of neck pain in the past decade. Before testing if there is an association between text neck and neck pain, it is necessary to develop reliable pragmatic tools appropriate to epidemiological studies.