Bone Marrow Neutrophil Storage Pool Research Articles

Colony stimulating factors as adjunctive therapy in neonatal sepsis

Colony stimulating factors as adjunctive therapy in neonatal sepsis

Immunological peculiarities of extremely preterm infants: a challenge for the prevention of nosocomial sepsis

To review the main aspects of fetal immune development focusing on the host defenses of extremely preterm infants against bacterial pathogens, and describing the possibilities of immunotherapeutic intervention for the prevention of neonatal nosocomial sepsis. Electronic search of MEDLINE database for articles published in the last 15 years. Those with relevant information regarding the target issue were selected. Immunity of extremely preterm infants is deficient due to skin fragility, insufficient complement system components, decreased bone marrow neutrophil storage pool, and lower chemotaxis, adherence, deformability, and neutrophil enzyme activities. Further limitations are found at NK cell-mediated cytotoxicity, T cell proliferation and cytokine production, B and T cell cooperation, and antibody synthesis by B lymphocytes. No definitive benefits of interventions for enhancing the immune function, such as the use of intravenous immunoglobulin or myeloid colony-stimulating factors, have been demonstrated. As a consequence of the immaturity of several immune components, extremely preterm infants are highly susceptible to nosocomial infections. The very limited possibilities for intervention in this system require the control of extrinsic factors for the prevention of nosocomial sepsis in these infants.

Hämatopoetische Wachstumsfaktoren in Prophylaxe und Therapie infektiöser Komplikationen bei Früh- und Neugeborenen1

Bacterial sepsis continues to be an important cause of morbidity and mortality in neonates. Neutropenia in the newborn, as a result of decreased bone marrow neutrophil storage pool reserves and myeloid committed progenitor cells, increases the risk of sepsis and is associated with a poor prognosis. The hematopoietic colony-stimulating factors G-CSF (granulocyte colony stimulating factor) and GM-CSF (granulocyte-macrophage colony stimulating factor) increase the number of circulating neutrophil number by stimulating neutrophil precursors proliferation. In a number of clinical trials in very low birthweight neonates and in neonates with preeclampsia-associated neonatal neutropenia, both hematopoietic growth factors significantly increased the circulating absolute neutrophil count. However, no larger study showed that prophylactic G-CSF or GM-CSF treatment resulted in a reduction of infectious complications or in an improved overall survival. Similar results were seen in studies evaluating G-CSF and GM-CSF as intervention therapy in septic neonates. Therefore, experts do not recommend the routine use of the expensive growth factors in preterm and term neonates. However, prospective clinical trials are still needed to evaluate whether specific treatment groups will benefit from the use of G-CSF or GM-CSF. In this regard, efforts must be directed at better defining the endpoints and in particular assigning value to reduction in treatment of possible infectious complications, such as days in hospital, antibiotic usage and costs. In addition, randomized studies are required to evaluate the proper dosage and duration of therapy, which most likely will vary between groups of patients.

Potential use of granulocyte colon-stimulating factor and granulocyte-macrophage colony-stimulating factor in neonates.

The immaturity of neonatal phagocytic immunity contributes to increased mortality during neonatal sepsis. Neonates have both quantitative and qualitative neutrophil defects with decreased bone marrow neutrophil storage pool (NSP) reserves, an inability to increase neutrophil production, and defective neutrophil functional activity. Neonates respond to overwhelming sepsis with depletion of the NSP and the development of peripheral neutropenia. The myelopoietic cytokines granulocyte colony-stimulating factor (G-CSF) and granulocyte-macrophage colony-stimulating factor (GM-CSF) have been documented to induce neutrophilia in neonatal animals and human infants, increase the NSP, and upregulate neutrophils for improved functional activity. Preclinical studies in neonatal rats demonstrate increased survival with prophylactic G-CSF during experimental group B streptococcal sepsis. In pilot phase I/II human trials, G-CSF and GM-CSF were demonstrated to be both safe and well tolerated and to induce significant increases in absolute neutrophil count and NSP. Prophylactic GM-CSF in the very low birth weight neonate may reduce the incidence of nosocomial infections. Phase III trials are needed to further delineate the clinical usefulness of these myelopoietic cytokines in neonates with a high predisposition to sepsis.

Results of a phase I/II trial of recombinant human granulocyte- macrophage colony-stimulating factor in very low birthweight neonates: significant induction of circulatory neutrophils, monocytes, platelets, and bone marrow neutrophils

Neonates, especially those of very low birthweight (VLBW), have an increased risk of nosocomial infections secondary to deficiencies in development. We previously demonstrated that granulocyte-macrophage colony-stimulating factor (GM-CSF) production and mRNA expression from stimulated neonatal mononuclear cells are significantly less than that from adult cells. Recombinant murine GM-CSF administration to neonatal rats has resulted in neutrophilia, increased neutrophil production, and increased survival of pups during experimental Staphylococcus aureus sepsis. In the present study, we sought to determine the safety and biologic response of recombinant human (rhu) GM-CSF in VLBW neonates. Twenty VLBW neonates (500 to 1,500 g), aged < 72 hours, were randomized to receive either placebo (n = 5) or rhuGM-CSF at 5.0 micrograms/kg once per day (n = 5), 5.0 micrograms/kg twice per day (n = 5), or 10 micrograms/kg once per day (n = 5) given via 2-hour intravenous infusion for 7 days. Complete blood counts, differential, and platelet counts were obtained, and tibial bone marrow aspirate was performed on day 8. Neutrophil C3bi receptor expression was measured at 0 and 24 hours. GM-CSF levels were measured by a sandwich enzyme-linked immunosorbent assay at 2, 4, 6, 12, and 24 hours after the first dose of rhuGM-CSF. At all doses, rhuGM-CSF was well tolerated, and there was no evidence of grade III or IV toxicity. Within 48 hours of administration, there was a significant increase in the circulating absolute neutrophil count (ANC) at 5.0 micrograms/kg twice per day and 10.0 micrograms/kg once per day, which continued for at least 24 hours after discontinuation of rhuGM-CSF. When the ANC was normalized for each patient's first ANC, there was a significant increase in the ANC on days 6 and 7 at each dose level. By day 7, all tested doses of rhuGM- CSF resulted in an increase in the absolute monocyte count (AMC) compared with placebo-treated neonates. In those receiving rhuGM-CSF 5.0 micrograms/kg twice per day, there was additionally a significant increase in the day 7 and 8 platelet count. Tibial bone marrow aspirates demonstrated a significant increase in the bone marrow neutrophil storage pool (BM NSP) at 5.0 micrograms/kg twice per day and 10.0 micrograms/kg once per day. Neutrophil C3bi receptor expression was significantly increased 24 hours after the first dose of rhuGM-CSF at 5.0 micrograms/kg once per day. The elimination half-life (T1/2) of rhuGM-CSF was 1.4 +/- 0.8 to 3.9 +/- 2.8 hours.(ABSTRACT TRUNCATED AT 400 WORDS)

Results of a Phase I/II Trial of Recombinant Human Granulocyte-Macrophage Colony-Stimulating Factor in Very Low Birthweight Neonates: Significant Induction of Circulatory Neutrophils, Monocytes, Platelets, and Bone Marrow Neutrophils

Neonates, especially those of very low birthweight (VLBW), have an increased risk of nosocomial infections secondary to deficiencies in development. We previously demonstrated that granulocyte-macrophage colony-stimulating factor (GM-CSF) production and mRNA expression from stimulated neonatal mononuclear cells are significantly less than that from adult cells. Recombinant murine GM-CSF administration to neonatal rats has resulted in neutrophilia, increased neutrophil production, and increased survival of pups during experimental Staphylococcus aureus sepsis. In the present study, we sought to determine the safety and biologic response of recombinant human (rhu) GM-CSF in VLBW neonates. Twenty VLBW neonates (500 to 1,500 g), aged < 72 hours, were randomized to receive either placebo (n = 5) or rhuGM-CSF at 5.0 micrograms/kg once per day (n = 5), 5.0 micrograms/kg twice per day (n = 5), or 10 micrograms/kg once per day (n = 5) given via 2-hour intravenous infusion for 7 days. Complete blood counts, differential, and platelet counts were obtained, and tibial bone marrow aspirate was performed on day 8. Neutrophil C3bi receptor expression was measured at 0 and 24 hours. GM-CSF levels were measured by a sandwich enzyme-linked immunosorbent assay at 2, 4, 6, 12, and 24 hours after the first dose of rhuGM-CSF. At all doses, rhuGM-CSF was well tolerated, and there was no evidence of grade III or IV toxicity. Within 48 hours of administration, there was a significant increase in the circulating absolute neutrophil count (ANC) at 5.0 micrograms/kg twice per day and 10.0 micrograms/kg once per day, which continued for at least 24 hours after discontinuation of rhuGM-CSF. When the ANC was normalized for each patient's first ANC, there was a significant increase in the ANC on days 6 and 7 at each dose level. By day 7, all tested doses of rhuGM-CSF resulted in an increase in the absolute monocyte count (AMC) compared with placebo-treated neonates. In those receiving rhuGM-CSF 5.0 micrograms/kg twice per day, there was additionally a significant increase in the day 7 and 8 platelet count. Tibial bone marrow aspirates demonstrated a significant increase in the bone marrow neutrophil storage pool (BM NSP) at 5.0 micrograms/kg twice per day and 10.0 micrograms/kg once per day. Neutrophil C3bi receptor expression was significantly increased 24 hours after the first dose of rhuGM-CSF at 5.0 micrograms/kg once per day. The elimination half-life (T1/2) of rhuGM-CSF was 1.4 +/- 0.8 to 3.9 +/- 2.8 hours.(ABSTRACT TRUNCATED AT 400 WORDS)

Granulocyte Colony-Stimulating Factor for Neutropenia in Neonatal Sepsis-Reply

We appreciate Nankervis and Seguin's patient report of another neonate who apparently benefited from rhG-CSF administration during bacterial sepsis. They aptly note the main differences in the two cases, namely route and duration of cytokine administration and neutrophil count response. Each patient had a very rapid rise in the neutrophil count, likely due to the release of mature neutrophils from the bone marrow neutrophil storage pool. 1 The differences in the duration of neutrophilia in the two cases may have reflected the subcutaneous vs intravenous routes of administration (prolonged systemic uptake from a subcutaneous drug depot in our case), the total number of doses given, or both. We disagree, however, that the single granulocyte transfusion that our patient received altered the degree of neutrophilia or natural outcome of his rhG-CSF response, since transfused granulocytes have such a short life span. 2,3 While both patients received aggressive supportive care that included

A randomized, placebo-controlled trial of recombinant human granulocyte colony-stimulating factor administration in newborn infants with presumed sepsis: significant induction of peripheral and bone marrow neutrophilia

Host defenses in the human neonate are limited by immaturity in phagocytic immunity. Such limitations seem to predispose infected newborns to neutropenia from an exhaustion of the neutrophil reserve. Among the critical defects thus far identified in neonatal phagocytic immunity is a specific reduction in the capacity of mononuclear cells to express granulocyte colony-stimulating factor (G-CSF) after stimulation. However, the safety, pharmacokinetics, and biological efficacy of administration of recombinant human (rh)G-CSF to infected human newborns to compensate for this deficiency is unknown. Forty-two newborn infants (26 to 40 weeks of age) with presumed bacterial sepsis within the first 3 days of life were randomized to receive either placebo or varying doses of rhG-CSF (1.0, 5.0 or 10.0 micrograms/kg every 24 hours [36 patients] or 5.0 or 10.0 micrograms/kg every 12 hours [6 patients]) on days 1, 2, and 3. Complete blood counts with differential and platelet counts were obtained at hours 0, 2, 6, 24, 48, 72, and 96. Circulating G-CSF concentrations were determined at hours 0, 2, 6, 12, 14, 16, 18, 24, and 36. Tibial bone marrow aspirates were obtained after 72 hours for quantification of the bone marrow neutrophil storage pool (NSP), neutrophil proliferative pool, granulocyte progenitors, and pluripotent progenitors. Functional activation of neutrophils (C3bi expression) was determined 24 hours after rhG-CSF or placebo administration. Intravenous rhG-CSF was not associated with any recognized acute toxicity. RhG-CSF induced a significant increase in the blood neutrophil concentration 24 hours after the 5 and 10 micrograms/kg doses every 12 and 24 hours and it was sustained as long as 96 hours. A dose-dependent increase in the NSP was seen following rhG-CSF. Neutrophil C3bi expression was significantly increased at 24 hours after 10 micrograms/kg every 24-hour dose of rhG- CSF. The half-life of rhG-CSF was 4.4 +/- 0.4 hours. The rhG-CSF was well tolerated at all gestational ages treated. The rhG-CSF induced a significant increase in the peripheral blood and bone marrow absolute neutrophil concentration and in C3bi expression. Future clinical trials aimed at improving the outcome of overwhelming bacterial sepsis and neutropenia in newborn infants might include the use of rhG-CSF.

A Randomized, Placebo-Controlled Trial of Recombinant HumanGranulocyte Colony-Stimulating Factor Administration in Newborn Infants With Presumed Sepsis: Significant Induction of Peripheral and Bone Marrow Neutrophilia

Host defenses in the human neonate are limited by immaturity in phagocytic immunity. Such limitations seem to predispose infected newborns to neutropenia from an exhaustion of the neutrophil reserve. Among the critical defects thus far identified in neonatal phagocytic immunity is a specific reduction in the capacity of mononuclear cells to express granulocyte colony-stimulating factor (G-CSF) after stimulation. However, the safety, pharmacokinetics, and biological efficacy of administration of recombinant human (rh)G-CSF to infected human newborns to compensate for this deficiency is unknown. Forty-two newborn infants (26 to 40 weeks of age) with presumed bacterial sepsis within the first 3 days of life were randomized to receive either placebo or varying doses of rhG-CSF (1.0, 5.0 or 10.0 micrograms/kg every 24 hours [36 patients] or 5.0 or 10.0 micrograms/kg every 12 hours [6 patients]) on days 1, 2, and 3. Complete blood counts with differential and platelet counts were obtained at hours 0, 2, 6, 24, 48, 72, and 96. Circulating G-CSF concentrations were determined at hours 0, 2, 6, 12, 14, 16, 18, 24, and 36. Tibial bone marrow aspirates were obtained after 72 hours for quantification of the bone marrow neutrophil storage pool (NSP), neutrophil proliferative pool, granulocyte progenitors, and pluripotent progenitors. Functional activation of neutrophils (C3bi expression) was determined 24 hours after rhG-CSF or placebo administration. Intravenous rhG-CSF was not associated with any recognized acute toxicity. RhG-CSF induced a significant increase in the blood neutrophil concentration 24 hours after the 5 and 10 micrograms/kg doses every 12 and 24 hours and it was sustained as long as 96 hours. A dose-dependent increase in the NSP was seen following rhG-CSF. Neutrophil C3bi expression was significantly increased at 24 hours after 10 micrograms/kg every 24-hour dose of rhG-CSF. The half-life of rhG-CSF was 4.4 +/- 0.4 hours. The rhG-CSF was well tolerated at all gestational ages treated. The rhG-CSF induced a significant increase in the peripheral blood and bone marrow absolute neutrophil concentration and in C3bi expression. Future clinical trials aimed at improving the outcome of overwhelming bacterial sepsis and neutropenia in newborn infants might include the use of rhG-CSF.

Randomized trial of granulocyte transfusions versus intravenous immune globulin therapy for neonatal neutropenia and sepsis

We prospectively studied newborn infants with sepsis and neutropenia who were randomly selected to receive standard supportive care and either adjuvant granulocyte transfusions or intravenous immune globulin (IVIG) infusions; 21 infants received granulocyte transfusions and 14 received IVIG infusions. Half of the patients were premature (gestational age less than or equal to 32 weeks); the average postnatal age was 5 days (range 3 to 8 days). All infants had neutropenia by the criteria of Manroe et al., and the mean average bone marrow neutrophil storage pool ranged between 35% and 37%. There were no significant differences with respect to serum IgG, IgA, IgM, and total hemolytic complement values between treatment groups or between survivors and nonsurvivors. Clinical severity as defined by hypoxia, acidosis, and hypotension was similar between treatment groups. Group B streptococcus was the most common organism identified and accounted for almost 33% of all bacterial isolates. There was a significantly different survival rate in the group receiving polymorphonuclear leukocyte transfusions (100%, 21/21) compared with the group receiving IVIG infusions (64%, 9/14; p = less than 0.03). There were no significant complications in either treatment group with respect to fluid overload, secondary infection, blood group sensitization, pulmonary complications, or graft-versus-host disease. This pilot study suggests a possible benefit of granulocyte transfusions compared with 'IVIG therapy in the adjuvant treatment of neonatal neutropenia and overwhelming bacterial sepsis.

Sequential Administration of Interleukin-6 and Granulocyte-Colony Stimulating Factor in Newborn Rats: Modulation of Newborn Granulopoiesis and Thrombopoiesis

During states of increased demand, neonatal host defense is characterized by dysregulation of granulopoiesis, resulting in a high incidence of neutropenia. This study investigated the modulation of neonatal rat hematopoiesis by 14-d administration of recombinant human (rh) IL-6, rh-granulocyte-colony stimulating factor (G-CSF), or sequential combination of rhIL-6 and rhG-CSF. Specifically, newborn Sprague-Dawley rats were treated with either rhIL-6 (5 micrograms/kg/d for 14 d), rhG-CSF (5 micrograms/kg/d for 14 d), rhIL-6 for 7 d followed by rhG-CSF for 7 d, PBS/BSA for 7 d followed by rhG-CSF for 7 d, or PBS/BSA for 14 d. RhIL-6 alone significantly increased the peripheral platelet count during the latter part of the 2nd wk of administration (d 13: 980 +/- 42 versus 716 +/- 23 x 10(3)/mm3) (p = less than 0.001) (mean +/- SEM). Treatment with rhIL-6 for 7 d followed by rhG-CSF significantly increased the peripheral neutrophil count compared with 7 d of PBS/BSA and 7 d of G-CSF (d 14 absolute neutrophil count 4888 +/- 12 versus 2720 +/- 317/mm3) (p = less than 0.05). Similarly, sequential rhIL-6/rhG-CSF significantly increased the d-14 bone marrow neutrophil storage pool (9873 +/- 882 versus 3564 +/- 159/mm3) (p = less than 0.005). Lastly, sequential rhIL-6/rhG-CSF induced the highest increase in bone marrow (p less than 0.01) and liver/spleen CFU-GM pool (p less than 0.001) compared with any other treatment group. These studies suggest that rhIL-6 alone is associated with a significant increase in the neonatal platelet count.(ABSTRACT TRUNCATED AT 250 WORDS)

Modulation of Neonatal Myelopoiesis in Newborn Rats after 7 Days' Administration of Either Granulocyte-Monocyte Colony Stimulating Factor or Interleukin-3

Single-pulse administration of either recombinant human granulocyte-monocyte colony stimulating factor or recombinant human granulocyte colony stimulating factor to newborn rats has previously been demonstrated to increase the peripheral neutrophil count and modulate bone marrow (BM) neutrophil pools. In our present study, we investigated the effects of 7 d of either recombinant murine granulocyte-monocyte colony stimulating factor (rmGM-CSF) (75 micrograms/kg/d) or recombinant murine IL-3 (rm IL-3) (10 micrograms/kg/d) on newborn rat myelopoiesis. Sprague Dawley newborn rats (greater than or equal to 24 h) were injected (intraperitoneally) daily for 7 d with either rmGM-CSF, rmIL-3, or PBS/BSA. rmGM-CSF induced a significant increase in the peripheral neutrophil count on d 3 (p less than 0.03) and d 7 (p less than 0.001) (75% increase). Additionally, rmGM-CSF induced a 50% increase in the BM neutrophil storage pool (p less than 0.025). rmIL-3 increased the BM colony forming unit-granulocyte monocyte pool (p less than 0.001); however, it failed to increase the peripheral neutrophil count or BM neutrophil storage pool. Neither CSF increased the BM neutrophil proliferative pool or BM colony forming unit-granulocyte monocyte proliferative rate. Additionally, 7 d of rmGM-CSF with or without antibiotics did not synergistically alter the mortality rate after group B streptococcol inoculation. This study suggests that rmIL-3 appears to stimulate more neonatal myeloid committed progenitor cell activity compared with rmGM-CSF. Optimal modulation of neonatal myelopoiesis may require the use of a sequential combination of hematopoietic CSF, namely an early-acting CSF followed by a more lineage myeloid-specific CSF.

The role of granulocyte transfusions as adjuvant therapy in the treatment of neonatal sepsis

Newborn host defense is characterized by an immaturity in phagocytic immunity and thereby predisposes the newborn to an increased incidence of overwhelming bacterial sepsis. Both qualitative and quantitative myeloid dysfunction are characteristic of neonatal host defense. Animal studies have suggested a possible role of granulocyte transfusions to immunomodulate neonatal host defense and reduce the high mortality associated with bacterial sepsis. Recently, we have demonstrated that granulocyte transfusions improve the survival rate of newborns with overwhelming bacterial sepsis. Adult donor granulocytes, 15 mL/kg/day twice a day for 2 days and once a day on day 3, administered to 31 septic newborns, were associated with a 97% survival rate compared to supportive care alone with or without IVIG (1000 mg/kg/day × 3 days) (52%) ( P < 0.004). A decrease in the bone marrow neutrophil storage pool (PMNs + bands + metamyelocytes) at the time of sepsis tends to be associated with a poor prognosis. Additional studies using granulocytes obtained by leukophereses for treatment of neonatal sepsis have demonstrated a similar improvement in survival, however, studies utilizing buffy coat transfusions have showed no difference. Additionally, adverse reactions to granulocyte transfusions in this population have been minimal to date. A multicenter trial is needed to fully evaluate the future role of granulocyte transfusions in this clinical setting. Alternatively, other methods to improve the qualitative and quantitative phagocytic defect in the newborn, such as the use of colony-stimulating factors, need to be explored.

Lymphokines: Enhancement by Granulocyte-Macrophage and Granulocyte Colony-Stimulating Factors of Neonatal Myeloid Kinetics and Functional Activation of Polymorphonuclear Leukocytes

Colony-stimulating factors, such as the granulocyte-macrophage and the granulocyte colony-stimulating factors (GM-CSF and G-CSF), are glycoproteins with biologic specificity defined by their ability to support proliferation and differentiation of hematopoietic cells of various lineages. Their physiologic activities include stimulation and proliferation of early stem cell precursors and functional activation of mature peripheral effector cells. Recently produced recombinant human (rh) GM-CSF and G-CSF have been demonstrated to regulate hematopoietic neutrophil progenitor colony growth; to stimulate the release of bone marrow neutrophil storage pools; and to prime mature effector functions, including chemotaxis, oxidative metabolism, phagocytosis, C3bi receptor expression, and antibody-dependent cytotoxicity in adults. We examined the effects of rh-GM-CSF on priming superoxide release and chemotaxis of neonatal (cord) polymorphonuclear leukocytes (PMNs) and of rh-G-CSF and rh-GM-CSF on bone marrow neutrophil egress in the neonatal rat. A time-response evaluation of the effect of rh-GM-CSF revealed enhanced release of superoxide by PMNs. PMN chemotaxis also was enhanced by rh-GM-CSF, with a maximal response occurring earlier than enhanced superoxide release. Intraperitoneal administration of rh-G-CSF or rh-GM-CSF to 1-day-old rats resulted in significant increases in white blood cell counts and significant early neutrophilia. Bone marrow examination revealed that the neutrophilia was secondary to egress and mild depletion of the neutrophil storage pool but that the neutrophil storage pool later returned to normal. These preliminary studies suggest that rh-GM-CSF and rh-G-CSF prime neonatal effector function and induces significant PMN egress and neutrophilia.(ABSTRACT TRUNCATED AT 250 WORDS)

1535 A CONTROLLED TRIAL OF GRANULOCYTE TRANSFUSIONS IN NEUTROPENIC NEONATES

To determine if granulocyte transfusions(Grtx) improve survival of neutropenic infants, we prospectively randomized 25 critically ill infants to +Grtx and -Grtx groups, matching for BW <1500 gm(n=8) or >1500gm(n=6) or necrotizing enterocolitis(NEC) (n=11). All infants had 2 absolute neutrophil counts(ANC) <1500 at presentation. Bone marrow neutrophil storage pools(NSP) were determined in 23/25. Grtx(0.1-0.9×109Gr/kg) were given daily until ANC > 1500, yet only 5 infants with NEC needed >1 Grtx. Blood CSF or peritoneal cultures were positive in 17/25 infants. 6/9 +Grtx and 9/12 -Grtx infants with a peripheral immature: total(I:T) granulocyte ratio ≥.8 survived. 1 +Grtx and 4 -Grtx infants with bone marrow NSP depletion ≤7% survived. 2 infants had a decreased PaO2 after a 3rd Grtx without X-ray changes. Neither an I:T ≥.8 nor an NSP ≤7% predicted survival. Also, Grtx did not improve the survival of the total 25 infants or the 17 infants with positive cultures. 4/9 deaths related to neurologic compromise, while 4/16 survivors have permanent neurologic deficits. In addition, of the 6 +Grtx infants with NEC, 2/5 perforations occurred after transfusion, while 4 infants developed strictures and 5, short gut syndrome. We conclude: l)neither an I:T ≥0.8 nor an NSP ≤7% predict mortality, and 2)Grtx do not improve survival. Repeated Grtx may increase the risk of respiratory compromise.