INTRODUCTION Early onset sepsis is difficult to diagnose due to nonspecific symptoms and a lack of
reliable tests. It can progress quickly, and lead to neurodevelopmental consequences or be fatal if not
treated. However, approximately 10-fold more newborns are treated with antibiotics empirically and often
unnecessarily. This study aimed to compare the management recommendations of the Neonatal Early
Onset Sepsis Calculator with those of the Centers for Disease Control/American Academy of Pediatrics
guidelines.

METHODS Neonatal Early Onset Sepsis Calculator was applied to the data set to examine how an alternative
model would perform compared to current guidelines published by the CDC and compared to current
practice within the institution. Chi square and kappa value agreement was used to determine the difference
between treatment recommendations of NEOS calculator and AAP guideline.

RESULTS Of the 330 patients who received therapy, only 14.2% were recommended empiric antibiotics by
the EOS calculator, compared to the 39% recommended by the CDC guidelines (p < 0.001, ? = 0.372).
Eleven patients were identified to have culture-positive sepsis.

CONCLUSION The number of infants suspected with EOS and subsequently require antibiotic use at birth
may be dramatically reduced with the use of the neonatal EOS calculator.

Human ; Sepsis ; Blood Culture

Abstract: The diagnosis of bacteremia relies on the isolation and identification of the bacteria from blood cultures, whether they are community-acquired or nosocomial in origin. However, studies have shown that, in the Philippines alone, physicians have been found to underutilize these laboratory examinations. Objectives: The goal of this study was to determine the influence of positive blood cultures and sensitivity test results on the antibiotic choices of pediatrics residents at the University of the Philippines – Philippine General Hospital (UP-PGH). Methods: A chart review of patients with positive blood cultures, who were 18 years old and below, and admitted initially at the UP-PGH Pediatric Emergency Room (UP-PGH PER) from August 1, 2004 to July 31, 2005 was performed. Excluded were patients who died before the release of the blood culture reports or discharged per request or against medical advice, post-operative patients, patients with presumed polymicrobial sepsis, and patients with contaminated blood cultures. Results: One hundred twenty two (122) patients with positive blood cultures were included: 87 or 71.3% of the isolates were community-acquired, the most common pathogens of which were gram-positive bacteria, Staphylococcus epidermidis (18.3%), followed by gram-negative Salmonella (11.5%). Among the patients diagnosed with bacteremia at the UP-PGH PER, Staph. epidermidis was also the most common pathogen; with 34% of all isolates acquired nosocomially. Other significant isolates included Pseudomonas putida, Pseudomonas aeruginosa, and Klebsiella sp. Prior to the release of the blood culture and sensitivity results, 45 of the 122 patients were already discharged. Therapy at the time of discharge was of questionable efficacy, accounting to 73.3%. Of the 77 patients discharged after the release of blood culture and sensitivity (CS) results, only 21(27%) of the antibiotic therapies were modified, and 56 (73%) were not modified at all. It is imperative to know, however, that 50% of the antibiotic therapies were modified a day after the corresponding blood culture and sensitivity (CS) results came out for patients who presented with nosocomial infection. Conclusion In general, blood culture and sensitivity test results have a limited effect on the antibiotic choices of pediatric residents at the UP-PGH (University of the Philippines – Philippine General Hospital).
Bacteremia ; Blood Culture ; Anti-Bacterial Agents

Purpose: Long-term culture- initiating cells(LTC-IC) are stem cells that have the capacities of long-term engraftment and helping to establish hematopoietic microenvironment. For evaluation of the LTC-IC, we measured the counts and function with multidimentional flowcytometry in long-term culture media. METHODS: Samples were obtained from umbilical cord blood, leukapheresis products and bone marrow(BM). LTC-IC were counted with flowcytometric analysis using anti- CD34, anti-CD38, and anti-HLA-DR antibodies at 0, 5, and 8 weeks. Cell adhesion molecule related with stem cell were evaluated with flowcytometric analysis also using anti-VCAM-1(CD106) and anti-VLA-4(CD49d) at 0 and 8 weeks. RESULTS: The proportion of CD34+/CD38- cell from fractionated mononuclear cells at 0 week were 0.46%, 0.044%, and 0.038% for BM, leukapheresis products, and umbilical cord blood respectively and then rapidly decreased at 5 weeks, but still persisted at 8 weeks in all three groups. The proportion of CD34+/HLA-DR- cells was the same tendency to CD34+/CD38-. VCAM+ expression rate from fractionated CD34+ cells at 0 and 8 weeks were 67.3% and 40.2% for BM and 64.1% 44.2% for umbilical cord blood but it was very low 31.2% and 5.1% for leukapheresis products. VLA-4+ expression rate for fractionated CD34+ cells at 0 and 8 weeks were similar tendency to VCAM+ cells. CONCLUSION: This study suggest that the count of LTC-IC decreased with time but still persisted until 8 weeks. Umbilical cord blood including BM help to establish the hematopoietic microenvironments.
Antibodies ; Cell Adhesion ; Culture Media ; Fetal Blood ; Leukapheresis ; Stem Cells

No abstract available.
Allografts* ; Animals ; Blood Transfusion* ; Cyclosporine* ; Lymphocyte Culture Test, Mixed* ; Rats*

OBJECTIVECurrently, thrombocytopenia is typically observed in patients undergoing hematopioetic stem cell transplantation (HSCT), high-dose chemotherapy or irradiation. Severe thrombocytopenia can cause intestinal and intracranial hemorrhage. To transfuse ex vivo-expanded megakaryocytes (MK) into patients can reinforce the ability of platelet formation and shorten the time of platelet recovery. Therefore it is one of the effective approaches to reduce the danger. The purpose of the present study was to explore the differences in MK expansion between CD(34)(+) stem cells derived from umbilical cord blood (CB) and peripheral blood (PB) and to establish the most optimal culture system.

METHODSMononuclear cells were isolated by density gradient centrifugation over Ficoll-Hypaque gradient solution. CD(34)(+) cells were isolated by positive selection using an immunomagnetic separation system and the selected CD(34)(+) cells were seeded in Iscove's modified Dulbecco's medium (IMDM) supplemented with fetal calf serum (FCS) and certain kinds of cytokines. After 15 - 17 days of culture, the cells were counted and the content of CD(41)(+) cells was determined by using flow cytometry, and the number of megakaryocyte colony-forming unit (CFU-MK) was simultaneously measured.

RESULTSAfter the defined days of culture, the cytokine combination of thrombopoietin (TPO) + fetal liver tyrosine kinase ligand (FL) + IL-6 + IL-3 showed to be the most suitable for both PB and CB to obtain high numbers of MK, and to be better than any of the other three groups (P < 0.05). The CD(41)(+) cells from CB were expanded by193 +/- 25 fold on day 14, and those from PB were expanded by 131 +/- 18 fold on day 10. The number of CD(41)(+) cells from both CB and PB decreased.

CONCLUSIONFor PB and CB, the cytokine combination of TPO + FL + IL-6 + IL-3 is most suitable for obtaining large number of MK and is the best combination for ex vivo MK expansion. MKs from CB seemed to have higher proliferation potential than that from PB, and the optimal culture duration of MK from PB is shorter than that of MK from CB.

Antigens, CD34 ; Cell Culture Techniques ; methods ; Cell Proliferation ; Cell Separation ; Culture Media ; Fetal Blood ; cytology ; Flow Cytometry ; Humans ; Megakaryocytes ; cytology

Observation of microcirculation plays an important role on the basic research and clinical diagnosis. However, an observation as such on the anesthetized patient will cause stress reaction, thus it will affect normal physiological state and interfere experimental results. At present, a method adopting dorsal microcirculatory chamber (DMC) to do in vivo observation in an unanesthetized state can eliminate the influence of anesthesia. Based on the research reports and practical applications of this method abroad, we summarize, in here, the configuration, function, observation techniques; the application of DMC; and the research states of microcirculation observation.
Animals ; Back ; blood supply ; Blood Flow Velocity ; Diffusion Chambers, Culture ; Humans ; Microcirculation ; physiology ; Monitoring, Physiologic ; methods ; Skin ; blood supply

To optimize the culture environment and protocol of hematopoietic cells' expansion, avoiding the fluctuation caused by medium changing in stirred culture and concentration gradient in static culture, the hematopoietic cells from cord blood (CB) were cultured in a stirred bioreactor connected with a cell retention system, which is a gravity sedimentation settler designed for hematopoietic cell. Total cells expanded 11.5 and 18.6 fold respectively in the twice perfusion stirred cultures, in which CFU-Mix was expanded 23.2 and 20.4 fold, CFU-GM 13.9 fold and 21.5 fold, BFU-E 8.0 fold and 6.9 fold, CD34+ cells 17.1 fold and 15.4 fold. After 12-day culture, it was obtained that 1082 x 10(6) total cells, 6.31 x 10(6) CFU-GM, 6.2 x 10(6) CFU-Mix and 23 x 10(6) CD34+ cells from 267 x 10(6) CB mononuclear cells (MNC) in the first culture, and 1080 x 10(6) total cells, 4.65 x 10(6) CFU-GM, 11.0 x 10(6) CFU-Mix, and 25.0 x 10(6) CD34+ cells from 180 x 10(6) CB MNC. These two cultures met to the clinical scale. Due to the optimized dissolved oxygen (DO) and stable culture environment, the rate of stem/progenitor cells to total cells in the perfusion culture was higher than that in T-flask cell-retention feeding culture. But the cell growth was inhibited in the later phase of perfusion culture, when the cell density is high. The inhibition should be attribute to the high cell density itself. The perfusion culture environment in bioreactor with optimal DO and pH controlling is more favorable for stem/progenitor cells' maintenance and expansion, and the expanded cells' number has reached a clinical scale. But the high cell density in the later phase of perfusion culture caused inhibition to mature hematopoietic cell's growth.
Bioreactors ; Cell Culture Techniques ; methods ; Cells, Cultured ; Fetal Blood ; cytology ; Hematopoietic Stem Cells ; cytology ; Humans

This study was aimed to investigate a beneficial approach for resolving the deficiency of blood source, preventing the infection resulting from blood transfusion and overcoming the knotty match of patients with rare blood group by using massive expansion of erythroid cells from cord blood CD34(+) cells in vitro. The CD34(+) cells from human cord blood were cultured in serum-free medium supplemented with stem cell factor (SCF), interleukin-3 (IL-3) and erythropoietin (EPO) for 1 week, then expansion and differentiation of CD34(+) cells into erythroid cells were supported by co-culture with human mesenchymal stem cells (MSCs) derived from bone marrow for 2 weeks. The results indicated that after culture for 23 days, the expansion multiple of total cell number reached 2.52 x 10(5), and over 95% of these cells were erythroid cells as compared with less than 1% of myelomonocytic (CD14(+) or CD15(+)) cells and megakaryocytic (CD41(+)) cells. However, the culture system without MSC support was significantly disadvantaged both in expansion ability and ratio of erythroid cells when compared with MSC supporting system. It is concluded that the erythroid cells can be produced from CD34(+) cells in large scale by culturing in the system comprised of cytokine sets and MSC feeders, in which MSCs can support the proliferation and differentiation of erythroid cells.
Antigens, CD34 ; Cell Culture Techniques ; methods ; Cell Differentiation ; Fetal Blood ; cytology ; Hematopoietic Stem Cells ; cytology ; Humans

Vascular tissue engineering is a novel approach by which an ideal vascualr graft constructed in vitro that will not be obstructed for a long time without immunological reaction after implantation. This article reviewed the definition of the tissue engineering blood vessel (TEBV), cellular resourses, the selection of biocompatible materials, the devising methods and the research achievements. Furthermore, it also discussed the current problems of TEBV and looked forward to future clinical application.
Biocompatible Materials ; Bioprosthesis ; Blood Vessel Prosthesis ; Cell Culture Techniques ; Tissue Engineering ; methods

OBJECTIVETo explore a new method for in vitro expansion of human mesenchymal stem cells (MSC) by using platelet-rich plasma (PRP) in place of fetal calf serum (FCS).

METHODSBone marrow(BM) samples were obtained from the proximal femurs of patients with normal haematopoietic function undergoing total hip arthroplasty. 2 x 10(5)/cm2 BM nucleated cells were seeded in 25 cm2 flasks for MSC cultivation containing one of the 3 mediums: complete Dexter medium with 12.5% FBS and 12.5% horse serum (medium1), alpha-MEM with 10% FCS (medium2) and alpha-MEM with 5% PRP(medium3). At the same time, 1 x 10(6) nucleated BM cells and same amount of nucleated cells from iliac aspirate were seeded in 25 cm2 tissue flasks, colony forming unit-fibroblast (CFU-F) assay.

RESULTSCulture-expanded cells from proximal femurs presented a typical fibroblast-like morphology. Cells were positive for SH2 (CD 105), SH3 (CD73), Thy-1 (CD90), while negative for CD34 and CD45. On induction, these cells could differentiate into osteoblasts. A significantly higher proliferative capacity of MSCs expanded in medium3 was observed in comparison to those in mediuml or 2 without alteration of the phenotype and the differentiation property. CFU-F assays indicated that bone marrow from the proximal femoral contained significantly more CFU-F than that from iliac aspirate.

CONCLUSIONPlatelet-rich plasma can be used in place of FCS to provide a safer and more effective culture condition to expand MSCs for clinical purpose. The proximal femur BM cells can be obtained in hip surgeries.

Blood Platelets ; Cell Culture Techniques ; methods ; Cells, Cultured ; Humans ; Mesenchymal Stromal Cells ; cytology ; Plasma