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Inflammatory bowel disease (IBD) present in childhood in 15% to 25% of cases. The aim of therapy in children is not only to guarantee normal growth but also to prevent relapse and to maintain remission. Steroids are effective to induce remission; however, resistance, dependency, and irreversible side effects can develop. The aim of this study was to determine whether treatment with repeated infusions of autologous red blood cells (RBCs) loaded with dexamethasone 21-phosphate (Dex 21-P) is safe and allows maintenance of long-term remission in children with steroid-dependent Crohn disease (CD).

Inflammatory bowel disease (IBD) present in childhood in 15% to 25% of cases. The aim of therapy in children is not only to guarantee normal growth but also to prevent relapse and to maintain remission. Steroids are effective to induce remission; however, resistance, dependency, and irreversible side effects can develop. The aim of this study was to determine whether treatment with repeated infusions of autologous red blood cells (RBCs) loaded with dexamethasone 21-phosphate (Dex 21-P) is safe and allows maintenance of long-term remission in children with steroid-dependent Crohn disease (CD). Eighteen consecutive pediatric patients who met the inclusion criteria were admitted to the study. Infusions of autologous RBCs loaded with Dex 21-P were performed every 4 weeks; the mean duration of treatment was 24 months. At the beginning of treatment and after 6, 12, and 24 months, we performed clinical evaluation according to the Pediatric Crohn Disease Activity Index (pCDAI). Assessment of body mass indexamethasone and bone mineral density by means of computerized bone mineralometry-dual energy x-ray absorptiometry, endoscopic evaluation, and hematic morning cortisol determination were also performed. During treatment, the mean pCDAI significantly decreased (P< 0.05); 78% of patients discontinued steroids. Determination of morning cortisol showed suppression only on the first day after infusion, followed by normalization of values. Endoscopic findings showed remission in 44% of patients. None of the patients experienced serious side effects. These data suggest that repeated infusions of RBCs loaded with Dex 21-P can be safe and useful to maintain long-term remission in pediatric patients with moderately active CD.

Several studies have suggested that the oxidative modification of low-density lipoprotein (LDL) could play a key role in the early stages of atherosclerosis. The susceptibility of LDL to oxidation has been found to be greater in patients with coronary heart disease. Familial hypercholesterolaemia (FH) is a powerful clinical model in which to study the predictive role of LDL in atherogenesis. LDL-apheresis is a treatment that is able to decrease lipid levels in plasma. This study was aimed at investigating the reducing capacity of erythrocytes and the in vitro susceptibility to oxidation of LDL isolated from patients with homozygous, heterozygous and double-heterozygous FH, who were treated fortnightly with LDL-apheresis or left untreated. In 14 FH patients, at baseline and after a cycle of treatment, the susceptibility of LDL to oxidative modification was analysed by studying the kinetics of conjugate diene formation. Plasma hydroperoxides, polyunsaturated fatty acid content, LDL electrophoretic mobility on agarose, the titre of auto-antibodies against oxidized LDL and serum paraoxonase activity were also measured. Furthermore, in order to evaluate a potential relationship between LDL oxidation and redox status, erythrocyte GSH and ATP levels were determined in FH patients treated regularly or never treated previously by LDL-apheresis. Unlike in the control group, the oxidative status of LDL in all FH patients was modified by LDL-apheresis, as revealed by the higher negative charge and the increase in levels of hydroperoxides and antibodies against oxidized LDL in the plasma. Our findings suggest both an acute effect and a long-term effect of LDL-apheresis in FH patients treated with dextran sulphate cellulose apheresis. The acute effect of LDL-apheresis on the susceptibility to oxidation of plasma and LDL was demonstrated by significant decreases in plasma hydroperoxide content, total LDL concentration and polyunsaturated fatty acid content. The increased resistance of LDL to oxidation was shown by prolongation of the lag time (P < 0.05) in samples after a single cycle of treatment. The long-term effect of LDL-apheresis was demonstrated by the comparable values for lag phases (obtained from the kinetics of conjugate diene formation) in patients under active treatment and controls. Compared with healthy controls and untreated patients, the erythrocyte GSH content was significantly higher (P⩽ 0.001) in the treated group, suggesting the activation of reducing mechanisms.

Peripheral blood lymphocytes from eight Fanconi anemia (FA) patients, 14 FA heterozygotes, and nine normal subjects have been tested for their susceptibility to chromosomal breakage induction by diepoxybutane (DEB) and by two peroxides. In addition, the effect of five antioxidants was investigated in standard cultures and in cultures stressed either with DEB or with butylhydroperoxide (BHP) or with hydrogen peroxide (H2O2). DEB, BHP, and H2O2 dramatically increased the chromosomal breakage levels in homozygous and heterozygous FA cells. A partial correction of chromosomal instability was obtained by treating the patients' lymphocytes with antioxidants. A “protective” effect was also noted in the DEB or peroxide-stressed lymphocytes of patients and heterozygotes, grown in the presence of antioxidants.

Highly purified progenitors (including erythroid [BFU-E], granulo- monocytic [CFU-GM], multipotent [CFU-GEMM] progenitors, as well as multipotent progenitors with self-renewal capacity [CFU-B]) express high-affinity growth factor receptors (GFRs), with prevalent interleukin-3 receptors (IL-3Rs) (2,700/cell), a > or = 10-fold lower number of IL-6Rs (145/cell) and granulocyte-macrophage colony- stimulating factor receptors (GM-CSFRs) (300/cell), and a barely detectable level of erythropoietin (Ep) receptors (75/cell). Hematopoietic growth factor (HGF) dosages inducing peak clonogenetic effects are associated with partial/subtotal occupancy of the homologous HGF receptor (HGFR). Cross-reactivity between GFRs and heterologous GFs (including IL-6, IL-3, GM-CSF, Ep, and the kit ligand [KL]) was explored by competition experiments on purified progenitors with radiolabeled and excess cold HGFs at +4 degrees C. No cross- reaction was observed between IL-6R, IL-3R, EpR, and the heterologous GFs, whereas the GM-CSFR showed cross-reactivity with IL-3 and, to a lesser extent, KL. Modulation of GFRs was examined after 18 or 40 hours of incubation with GF(s) at 37 degrees C, followed by ligand-binding assay at 20 degrees C. IL-6, IL-3, GM-CSF, and Ep induce a marked down- modulation of their own receptors. Interestingly, each GF induces the transactivation of the R(s) for the “distal” GF(s): (1) IL-6 induces transactivation of IL-3R, but not of GM-CSFR/EpR; (2) IL-3 causes a rapid upmodulation of GM-CSFR/EpR (“pure” progenitors treated with IL-3 show upmodulation of GM-CSFR alpha-chain mRNA by reverse transcriptase- polymerase chain reaction); whereas (3) GM-CSF induces the transactivation of the EpR. This chain upmodulation of HGFRs may underlie the synergistic interactions between the HGFs in clonogenetic culture. It is emphasized that KL does not induce upmodulation of the other GFRs. Finally, Ep, GM-CSF, and IL-3 do not modulate the expression of the “proximal” HGFRs (ie, GM-CSFR/IL-3R/IL-6R, IL-3R/IL- 6R, and IL-6R, respectively). These results allow insight into the cellular basis of hematopoiesis, ie, the complex and coordinate interactions between HGFs and their receptors. They are compatible with a model of cascade transactivation via upmodulation of GFRs in the initial key steps of hematopoietic differentiation, whereby the action of each GF enhances the effect of the distal GF(s) by a multistep chain- potentiation mechanism.

In a 5-year-old Italian girl with severe congenital hemolytic anemia, red cell GPI deficiency was proven, and found to be due to a new variant, ‘GPI Roma.’ The parents are first cousins and have been proven to be heterozygous for this variant.

Summary We have developed a culture system for “longterm” growth of human lymphokine-activated killer (LAK) cells exhibiting an elevated, wide-spectrum antitumor cytotoxicity. The system allows the exponential growth of monocyte-depleted low-density lymphocytes in the presence of human serum and recombinant human interleukin-2 (103 U/ml), alone or in combination with interleukin-1 α or β (both at 10 U/ml). Eighteen cultures were established from 18 normal adult donors. The membrane phenotypes of the final LAK cell population, assessed by a panel of monoclonal antibodies (mAb), consist of three main types: (a) NKH-1+, Tiα/β−, Tiγ/δ−, and CD3− lymphocytes; (b) NKH-1+, Tiα/β−, Tiγ/δ+, and CD3+ lymphocytes and (c) NKH-1+, Tiα/β+, Tiγ/δ− and CD3+ lymphocytes. Northern blot analysis showed that all these cell populations express relatively high levels of perforin RNA, particularly cells exhibiting the first phenotype. This culture system may provide a tool for cellular and molecular studies on the mechanisms of antitumor cytotoxicity, as well as the basis for new adoptive immunotherapy protocols in advanced cancer.

Two sisters, double heterozygotes for Hb Lepore and β-thalassemia, of Italian origin (Naples), were extensively studied because of their particularly mild clinical and hematological picture, rather unusual in this type of syndrome. Family studies were carried out, and two Hb Lepore heterozygous carriers (the mother and a sister) and two β-thalassemia heterozygotes (the father and a brother) were identified. In vitro biosynthetic studies carried out in both a Hb Lepore carrier and the Hb Lepore/β-thalassemia double heterozygotes indicated an evident incorporation of radioactivity into the chromatographic peak which structural studies demonstrated to be Boston (= Washington) type of δβ-chain (i.e., δ87 Gln β116 His). The amounts of synthesis were roughly of the same order of magnitude as the relative levels of Hb Lepore in the red cell hemolysates, indicating the presence, in the reticulocytes of these subjects, of stable δβ mRNA molecules. These findings provide the opportunity to reconsider the current views indicating the instability of mRNAs as the cause of the low synthetic activity of the δβ and βδ fusion genes.

We have investigated the effect of iron on the expression of transferrin receptors (TrfRs) and ferritin chains in cultures of human peripheral blood monocytes maturing to macrophages.

At the end of 1960, the concept that platelet transfusion could reduce the death rate due to hemorrhage was confirmed by the reduction of life-threatening bleeding and prolonged survival in thrombocytopenic patients affected with acute leukemia and aplastic anemia. Thrombocytopenia is the result of an imbalance between platelet production and destruction: usually no bleeding problem will occur until circulating platelets fall below 20000/μl and even a platelet count of 5000/μl may be present in many patients without bleeding. Because of the high risk of alloimmunization in multiply transfused thrombocytopenic patients with random platelet concentrates, the main dilemma is the choice of strategy: “prophylactic”versus “therapeutic” treatment with platelet concentrates of cancer patients and the platelet count selected as the “transfusion trigger” for platelet support in patients without active bleeding. We describe our experience of 367 retrospective non-randomized leukemic patients transfused with platelet concentrates. A total of 225 patients (61.3%) received support therapy: the transfusions were administered prophylactically at a platelet count below 20000/μl in the group of patients with acute lymphoblastic leukemia (35% transfused) and acute myeloblastic leukemia (78% transfused). Only14 hemorrhagic episodes were observed in 148 patients receiving prophylactic platelets (9%), while 21 severe hemorrhages (27%) were documented in patients treated with therapeutic platelet concentrates. Several studies have concluded that maintaining the platelet count above 20000/μl was not justified in the majority of cancer patients. In the absence of more definitive data, a “transfusion trigger” of 10000/μl is selected for platelet transfusion support in leukemic non-bleeding patients receiving chemotherapy.

Central venous access is necessary in patients candidate for peripheral blood stem cell (PBSC) collection. We report our experience with a dual lumen femoral catheter (Gamcath, 11 french), initially designed for hemodialysis. We studied 147 patients and performed 488 collections after mobilization with either G-CSF alone or chemotherapy + G-CSF, when the white blood cell count exceeded 1 × 109/L, or when a measurable population of CD34+ cells (20 / μL) was detected in peripheral blood. All patients received systemic anticoagulation with a low weight heparin and ultrasound examination was performed after the removal of the catheter. Seven patients developed thrombosis (4.7%), ten experienced hematomas at the site of catether placement (6.8%) despite prophylactic platelet transfusions, while only one patient (0.6%) had a catheter-related infection. In conclusion, the short-term use of large bore femoral catheters in setting up PBSC collection seems to be associated with minimal risk of infection and low thrombotic incidence.

The action of the iron chelator desferrioxamine (DFO) on the cytogenetic pattern of cultured lymphocytes from Fanconi anemia (FA) patients was investigated. The addition of 10-4M DFO throughout the culture time resulted in a 50% reduction of the spontaneous chromosome breakage of FA cells. In addition, the clastogenic action of diepoxybutane on FA lymphocytes was also partly counteracted by DFO. The above findings support the assumption that one of the mechanisms involved in the pathogenesis of FA might be an impaired capacity of the cells from such patients to remove active oxygen species. The relationship between intraleukocyte chelatable iron pool and free radical formation in FA subjects is discussed.

In premature infants with sepsis several defects of PMN function have been documented, suggesting that these infants can be equated to neutropenic patients. We, therefore, evaluated the effectiveness of PMN transfusion in 11 very premature newborn infants with sepsis (b.w.820-1200 g; g.a. 25-29 wks). The diagnosis of sepsis was established according to clinical signs and to blood picture, and was always confirmed by blood cultures. Transfusion of packed PMNs (0.5 × 1010cells in 20 ml, obtained by leukafiltration of ABO/R h compatible blood) was given after sepsis was ascertained and repeated in the subsequent days. Group A (6 cases) received a mean of 1 unit per 2.2 symptomatic days, whereas Group B (5 cases) received a mean of 1 unit per 6 symptomatic days. The mortality rate was significantly lower in Group A than in Group B (17% vs. 100%; p <0.02). No side effect was observed. The present results suggest that PMN transfusion may represent a substantial improvement in the treatment of these infants. Adverse effect of the treatment needs further evaluation, but at present possible risks seem to be reasonably low. ( CNR grant no. 78.00563.83).

In the decade 1970-79, NEC was observed only in the period '76-'79, and only in infants weighing = 1250 g, with sepsis and surviving at least 48 hours. In 1978 a trial of PMNs transf. to septic infants was started. In septic infants with the above characteristics admitted in '76-'79, we considered the following items: sex, weight, GA, SBP, a-b status, temp. on adm., umb. vessels catheterization, feeding schedules, blood culture, clinical signs, antibiotic and PMNs transf. therapy, and occurrence of NEC. The 10 patients with and the 28 pts. without PMNs were similar for the above items, except that NEC occurred in 10/28 pts. without PMNs transf., and never in the 10 transfused pts. (p = 0.03). The 10 pts. with NEC and the 28 pts. without NEC were similar for the above items, except that PMN transf. had been given to 10/28 of the non-NEC pts., and never to the NEC pts. (p = 0.03). It is suggested that PMNs transfusion prevents the occurrence of NEC in septic very low birthweight infants.

25 newborns with bacterial sepsis received from 2 to 14 transfusions (20 ml/kg of a suspension containing .5-1 × 109PMNs, and less than 6% lymphocytes, obtained by continuous flow filtration leukapheresis). Results were compared with findings in 40 septic newborns not treated by PMNs transfusions (GROUP B). GROUPS A and B were comparable with respect to clinical characteristics and etiology (in most cases a highly antibiotic-resistant Klebsiella). The mortality rate was lower in GROUP A than in GROUP B (8% vs 70%, p <0.001). The same was true when comparing infants weighing <1500 g (8% vs 83%, p <0.001) or >1500 g (8% vs 50%, p = 0.02). Major complications and associated conditions (i.e. NEC, meningitis, pneumonia, peritonitis, osteoarthritis, DIC) were observed in 18 patients of GROUP B, and only in 3 of GROUP A. Untoward effects attributable to the PMNs transfusion were never observed. It is concluded that PMNs transfusion represents an highly effective treatment of neonatal sepsis. (C.N.R. project MPP3, n. 79.010.022.83).