Acute myocardial infarction after bone marrow transplantation: an unsuspected late complication.

Acute myocardial infarction is a common disease rarely seen as a complication of bone marrow transplantation in young patients. We report on a 25-year-old patient 3.5 years after bone marrow transplantation who suffered an acute anterior wall myocardial infarction complicated by cardiogenic shock. The patient was treated with thrombolysis and emergent coronary angioplasty but died a few hours following admission. We suggest that the combination of low-dose chest irradiation and prolonged immunosuppression with graft-versus-host disease contributed to the development of the coronary artery disease in this patient. Though rarely encountered, physicians caring for young patients after bone marrow transplantation should be aware of potential ischemic complications.

Molecular diagnosis of FMF: lessons from a study of 446 unrelated individuals.

BACKGROUND: Traditionally, the diagnosis of familial Mediterranean fever (FMF) has been based on clinical manifestations and the physician's experience. Following the cloning of the gene associated with this disease (MEFV), genetic analysis of its mutations has become available, providing a new tool for the establishment or confirmation of the diagnosis of FMF. OBJECTIVES: We analyzed the results of molecular testing for MEFV mutations in 600 individuals. We wished to determine how many of them bore mutations and what percentage had clinically active FMF. We also compared the rate of genetic confirmation of the FMF diagnosis in referrals with suspected FMF seen by general practitioners with that of persons sent for genetic analysis by FMF experts. METHODS: Of 600 individuals tested for FMF mutations, we analyzed separately 446 unrelated persons for the combination of their mutations, epidemiological data, and clinical manifestations. The five most common mutations in the present cohort were analyzed using the amplification refractory mutation system (ARMS). RESULTS: Of the 446 subjects analyzed, 249 (55%) bore mutations: 147 of these were homozygotes or compound heterozygotes, all of whom had FMF according to clinical criteria. Of the remaining 102 heterozygotes, 72 had FMF according to clinical criteria. Two patients with none of the five mutations also had FMF: North African Jews bore mainly mutations M694V and E148Q. The M6941 mutation was found exclusively in Palestinian Arabs. The rate of confirmation of FMF diagnosis by mutation analysis in subjects sent by FMF experts was significantly higher than that of persons referred by general practitioners. Analysis of the molecular testing of the multicase families (154 individuals) revealed that 141 of them bore MEFV mutations and that 4 persons homozygous for E148Q were asymptomatic. CONCLUSIONS: Molecular analysis of FMF mutations confirmed the diagnosis in about 60% of the referrals with suspected FMF. Some (33%) of the patients were heterozygotes, and there were also FMF patients with none of the 5 mutations analyzed. A second opinion by an FMF expert may decrease the need for mutation analysis in subjects suspected of having FMF.

Effect of ozone on neutrophil function in vitro.

The application of ozone is widely practised as a form of alternative medicine, particularly in Germany and Eastern Europe. Ozone major autohemotherapy (the return of a small amount of a patient's blood to the circulation after ex vivo exposure to ozone) has been reported to have a therapeutic effect in various pathological conditions, including ischemic, infectious, autoimmune and neoplastic disorders. Ozone has an effect on the expression of cytokines, adhesion molecules and acute phase reactants, which are responsible in part for the respiratory inflammatory response observed after exposure to this gas. The purpose of the present study was to investigate the effect of ozone administration ex vivo, at a concentration commonly used in major autohemotherapy, on peripheral blood neutrophil function in vitro. Blood drawn from healthy volunteers was studied for neutrophil adhesion, chemotaxis and O-2 production before and after exposure to 30 microg/ml ozone. There was no significant difference in adhesion and chemotaxis of neutrophils exposed to ozone versus unexposed cells. O-2 production was minimally decreased (20.3 +/- 5.0 vs. 22.1 +/- 5.5 nmol/106 cells/10 min, respectively; P=0.01), a reduction of no clinical significance. This study confirms that major autohemotherapy with ozone is safe as far as neutrophil function is concerned. Combined with previous data, it seems that well-designed clinical trials to assess the efficacy of major autohemotherapy would pose no danger to blood cell populations, and should be encouraged.

A novel computational method for predicting the transmembrane structure of G-protein coupled receptors: application to human C5aR and C3aR.

A novel algorithm was applied to the sequences of bacteriorhodopsin (BRh), of rhodopsin (Rh), and of the two human anaphylatoxin receptors, C5a-receptor (hC5aR) and C3a-receptor (hC3aR), that predicts their transmembrane domains (TMD) according to energy criteria alone, on the basis of their sequences and a template structure for each. Two consecutive criteria were applied for the predictions: the first is hydrophobicity of a sequence of residues, which determines the candidate stretches of residues that form one of the transmembrane helices. The second criterion is an energy function composed of inter residue contact energies, of hydrophobic contributions due to membrane exposure and of the interactions of a few residues with the phospholipid head groups. The sequence of candidate residues for each helix is longer than that of the template, and is finally determined by threading each of the candidate stretches on each of the template helices and evaluating the energy for all possible configurations. Contact energies between residues were taken from a database (Miyazawa S and Jernigan RL (1996) J Mol Biol 256 623-44). The algorithm predicts well the TMD structure of BRh based on its own template, and the TMD structure of Rh conforms well with the model of Baldwin et al (Baldwin JM Schertler GFX and Unger VM (1997) J Biol Chem 272 144-64). Results for the construction of the TMD of hC5aR and hC3aR were compared, employing the template structure of Rh. Most of the results for these receptors are in accord with alignments and with mutation experiments on hC5aR and hC3aR. The predictions may serve as a basis for future mutagenesis experiments of these receptors.

Expression of the familial Mediterranean fever gene and activity of the C5a inhibitor in human primary fibroblast cultures.

Familial Mediterranean fever (FMF) is an inherited disease whose manifestations are acute but reversible attacks of sterile inflammation affecting synovial and serosal spaces. The FMF gene (MEFV) was recently cloned, and it codes for a protein (pyrin/marenostrin) homologous to known nuclear factors. We previously reported the deficient activity of a C5a/interleukin (IL)-8 inhibitor, a physiologic regulator of inflammatory processes, in FMF serosal and synovial fluids. We now describe the concomitant expression of MEFV and C5a/IL-8-inhibitor activity in primary cultures of human fibroblasts. Fibroblasts grown from synovial and peritoneal tissues displayed C5a/IL-8-inhibitor activity that could be further induced with phorbol myristate acetate (PMA) and IL-1 beta. Very low levels of chemotactic inhibitor were evident in skin fibroblast cultures or in peritoneal and skin fibroblasts obtained from FMF patients. MEFV was expressed in peritoneal and skin fibroblasts at a lower level than in neutrophils and could be further induced by PMA and IL-1 beta. In the FMF cultures, the MEFV transcript carried the M694V mutation, consistent with the genetic defect found in patients with this disease. MEFV was also expressed in other cell lines that do not produce C5a/IL-8 inhibitor. These findings suggest that human primary fibroblast cultures express MEFV and produce C5a/IL-8-inhibitor activity. The interrelationship between pyrin, the MEFV product, and the C5a/IL-8 inhibitor requires further investigation. (Blood. 2000;96:727-731)

Expression and function of serum amyloid A, a major acute-phase protein, in normal and disease states.

Serum amyloid A (SAA), the precursor protein in inflammation-associated reactive amyloidosis (AA-type), is an acute phase reactant whose level in the blood increases in response to various insults. It is expressed in the liver, but its physiological role is not well understood. Recently, a broader view of SAA expression and function has been emerging. Expression studies show local production of SAA proteins in histologically normal, atherosclerotic, Alzheimer, inflammatory, and tumor tissues. Binding sites in the SAA protein for high density lipoproteins, calcium, laminin, and heparin/heparan-sulfate were described. Adhesion motifs were identified and new functions, affecting cell adhesion, migration, proliferation and aggregation have been described. These findings emphasize the importance of SAA in various physiological and pathological processes, including inflammation, atherosclerosis, thrombosis, AA-amyloidosis, rheumatoid arthritis, and neoplasia. In addition, recent experiments suggest that SAA may play a "housekeeping" role in normal human tissues.

MEFV mutations in Turkish patients suffering from Familial Mediterranean Fever.

Familial Mediterranean fever (FMF) is a recessive inherited disorder affecting Sephardic Jews, Arabs, Armenians and Turks. The gene responsible for FMF was recently cloned and several disease-associated mutations have been described. We have evaluated seven MEFV mutations in 460 chromosomes of 230 unrelated patients with FMF living in Turkey, using PCR methods. The M694V allele accounted for 43.5% of the alleles studied and 19.1% of the patients were homozygous. The M680I, V726A and M694I mutations were responsible for 12.0%, 11.1% and 2.8% of the patients respectively. R761H, K695R and E148Q were rarely encountered. Two thirds of the disease alleles were attributed to three common mutations: M694V, M680V and V726A, but only 54% of the patients carried one or two of the three mutations. Adding the four rarer mutations increased these figures to 72% and 60%, respectively. Altogether, 79.6% of the patients bore at least one of the main mutations, and 84.3% carried at least one of the seven mutations studied. The 28 patients suffering also from amyloidosis carried at least one of five mutations, M694V being the most common. These results suggest that the origin of FMF in Turkey is heterogenous, all common mutations are associated with amyloidosis. Further, rapid and accurate molecular diagnosis of FMF is feasible in most cases.

Effect of serum amyloid A on selected in vitro functions of isolated human neutrophils.

Serum amyloid A (SAA) is an acute phase reactant whose levels in the blood rise as part of the body's response to stress and inflammation. Previous studies have suggested that SAA may carry an anti-inflammatory potential. We evaluated the effects of SAA on human neutrophils activated by N-formyl-methionyl-leucyl-phenylalanine (fMLP) in vitro. At concentrations higher than 10 microg/mL, SAA inhibited neutrophil myeloperoxidase (MPO) release. This effect was located in the N-terminal--that is, amino acid residues 1-14--of the SAA molecule. Directed neutrophil migration was inhibited at the same SAA concentrations. Several amino acid residues (1-14, 15-104, 83-104) contributed to this effect. Neutrophil O2- production was inhibited at low concentrations of SAA (0.1 to 1 microg/ml) and was stimulated at concentrations higher than 50 microg/mL. Neutrophil O2- production induced by phorbol myristate acetate (PMA) and O2- generated by the xanthine-xanthine oxidase reaction were not affected by SAA. These results add to previous data suggesting that SAA, at concentrations recorded in the serum during inflammation, modulates neutrophil function; thus it may play a role in the down-regulation of the inflammatory process.

Widespread expression of serum amyloid A in histologically normal human tissues. Predominant localization to the epithelium.

Serum amyloid A (SAA) is an acute-phase reactant whose level in the blood is elevated to 1000-fold as part of the body's responses to various injuries, including trauma, infection, inflammation, and neoplasia. As an acute-phase reactant, the liver has been considered to be the primary site of expression. However, limited extrahepatic SAA expression was described in mouse tissues and in cells of human atherosclerotic lesions. Here we describe nonradioactive in situ hybridization experiments revealing that the SAA mRNA is widely expressed in many histologically normal human tissues. Expression was localized predominantly to the epithelial components of a variety of tissues, including breast, stomach, small and large intestine, prostate, lung, pancreas, kidney, tonsil, thyroid, pituitary, placenta, skin epidermis, and brain neurons. Expression was also observed in lymphocytes, plasma cells, and endothelial cells. RT-PCR analysis of selected tissues revealed expression of the SAA1, SAA2, and SAA4 genes but not of SAA3, consistent with expression of these genes in the liver. Immunohistochemical staining revealed SAA protein expression that co-localized with SAA mRNA expression. These data indicate local production of the SAA proteins in histologically normal human extrahepatic tissues.

Diagnosis of familial Mediterranean fever by a molecular genetics method.

BACKGROUND: Familial Mediterranean fever is a recessively inherited disorder characterized by episodes of fever with abdominal pain, pleurisy, or arthritis. The familial Mediterranean fever gene, designated MEFV, was recently cloned, and at least three missense mutations (M6801, M694V, and V726A) that account for a large percentage of patients with this disease were identified. OBJECTIVE: To establish a diagnostic test for familial Mediterranean fever. DESIGN: Cross-sectional study of a convenience sample of patients attending familial Mediterranean fever clinics. SETTING: Tertiary referral hospitals. PATIENTS: 107 patients with familial Mediterranean fever, their family members, and controls. MEASUREMENTS: Mutations in the 107 samples were assessed by amplifying genomic DNA with use of primers that selectively amplify the normal or altered DNA sequence of the 3 MEFV mutations (amplification refractory mutation system [ARMS]). Mutations were independently assessed by automated sequencing of genomic DNA amplified by polymerase chain reaction to evaluate the sensitivity and specificity of the ARMS assay. RESULTS: The ARMS assay correctly identified M6801, M694V, and V726A mutations in 82 persons with mutations documented by DNA sequencing (21 homozygotes, 2 compound heterozygotes, and 59 simple heterozygotes). Of 7 persons known from family studies to be noncarriers and 18 unrelated persons who were negative for these mutations by sequencing, none had MEFV mutations according to ARMS. CONCLUSION: The ARMS assay is a rapid, cost-effective, and accurate method for detecting three common mutations in familial Mediterranean fever.

Anticardiolipin antibodies in acute myeloid leukemia: prevalence and clinical significance.

This prospective study was designed to explore the prevalence and the clinical and prognostic significance of anticardiolipin (ACL) antibodies in patients with acute myeloid leukemia (AML). The study includes 37 consecutive AML patients >15 years old without previous history of thromboembolism, recurrent fetal loss, or autoimmune disease and with no evidence of infection at the time of enrollment. ACL antibodies were found in 25 patients (68%). None of the patients had high positive titers; 8 had moderately positive while 17 had low positive ACL antibody titers. ACL antibody positivity did not predict response to chemotherapy and was not correlated with age, gender, FAB class, platelet and white blood cell counts at presentation, and the risk of thromboembolism. ACL antibody titers did correlate, however, with AML activity in the majority of patients (93%) during 4-19 months of follow up. These results demonstrate a high prevalence of ACL antibodies in AML patients and suggest that serum ACL antibodies may be a useful adjunct in predicting relapse and documenting disease activity and therapy response.

Combined clozapine-lithium treatment for schizophrenia and schizoaffective disorder.

Three adolescent and two adult patients suffering from chronic excited psychoses (either schizophrenia or schizoaffective disorder) resistant to traditional neuroleptics and clozapine were treated with combined clozapine-lithium. Improvement was assessed with the Positive and Negative Symptoms Scale, the Brief Psychiatric Rating Scale and the Clinical Global Impressions, administered before and during combined clozapine-lithium treatment. All patients demonstrated a significant improvement with this combination. There was no occurrence of agranulocytosis, neuroleptic malignant syndrome or other clinically significant adverse effects.

Effect of theophylline on erythrocytosis in chronic obstructive pulmonary disease.

BACKGROUND: Patients with chronic obstructive pulmonary disease (COPD) tend to develop secondary erythrocytosis to compensate for their chronic hypoxia. Theophylline has recently been shown to reduce hematocrit and erythropoietin blood levels in normal subjects and in patients with erythrocytosis after renal transplantation. OBJECTIVE: To determine whether theophylline may be used to lower the hematocrit in patients with COPD. METHODS: Two hundred four patients with COPD were studied retrospectively and 10 patients prospectively (8 starting treatment with the drug [group 1] and 2 who suspended its long-term use [group 2]) for the correlation between theophylline therapy and hematocrit and erythropoietin level. RESULTS: In the patients studied retrospectively, lower hematocrits were found in the theophylline-treated than in the untreated patients (0.43 +/- 0.006 vs 0.46 +/- 0.007, respectively; P < .002). Twelve untreated patients and 2 of those treated with theophylline had hematocrits above 52%. Oxygen saturation levels were similar in both groups, and exclusion of patients with oxygen saturation lower than 88% did not change the pattern, suggesting that the effect of theophylline could not be entirely explained by improved oxygen availability. Seven of the 8 patients studied prospectively in group 1 (P < .02) and the 2 patients in group 2 showed inverse correlations between hematocrits and theophylline administration. A similar pattern was observed with serum erythropoietin levels in 5 of 7 patients studied. The effects were reproducible on rechallenge in 3 of the 4 patients in group 1 and the 2 patients in group 2. CONCLUSIONS: Theophylline may have a beneficial effect in treatment and prevention of erythrocytosis in patients with COPD.

Laparoscopic splenectomy for immune thrombocytopenic purpura.

Splenectomy is an effective treatment for immune thrombocytopenic purpura (ITP). The recent advances in laparoscopic technique and technology have made laparoscopic splenectomy a viable option. Over 36 months we performed a total of 17 laparoscopic splenectomies, 15 of them for ITP and 2 for familial spherocytosis. We present our initial experience with laparoscopic splenectomy in 15 patients (age 16-71 years) with ITP. Operations were performed 2-24 months after the establishment of the diagnosis and initiation of appropriate therapy. Technically, the splenic artery was clipped first; the lower pole of the spleen and its posterolateral attachments were dissected using endoclips and electrocautery; the hilum and short gastric vessels were separated using an endostapler; the spleen was placed in a plastic bag, its opening pulled out through the umbilical incision, and the spleen fragmented and aspirated out of the bag. Operations lasted 100-300 min (mean 170 min). No patient required blood transfusion. The postoperative course was uneventful in all patients with minimal requirement of analgesia and early return to normal activity. Platelet counts returned to normal in all patients in a follow-up period of 2-36 months. Laparoscopic splenectomy is safe and effective for patients with ITP because of reduced operative trauma, less postoperative pain, cosmetic advantage, and possibly less postoperative complications.