Pro-inflammatory effector Th cells transmigrate through anti-inflammatory environments into the murine fetus.

The presence of maternal DNA or even maternal cells within the offspring (microchimerism) has been reported for many fetal tissues, including the liver, heart, and spleen. Microchimerism is believed to be involved in the pathogenesis of autoimmune diseases; however, the cellular origin of this phenomenon remains unknown. Here, we determined whether differentiated T lymphocytes could transmigrate through the immunosuppressive environment of the placenta to reach the fetus. In vitro-differentiated effector/memory Th1 and Th17 cells from OVA323₋339-specific TCR(tg) T cells of OT-II mice were adoptively transferred (i.v.) into the tail veins of pregnant Ly5.1 mice at d15 and d19 of gestation. Mice were then sacrificed 40 h after adoptive cell transfer. Using radioactive labeling of T cells with sodium chromate [Cr5¹] prior to adoptive transfer, we observed that homing of pro-inflammatory Th cells was equally efficient in both pregnant and non-pregnant mice. Transmigration of Th1- and Th17-like cells through the highly immunosuppressive environment of the placenta into the fetus was significantly enhanced in experimental mice compared to control mice (P < 0.0001). In addition, a substantial amount of effector Th cells accumulated in the placenta. Finally, we found that treatment with Pertussis Toxin resulted in a 3-fold increase in the transmigration of effector Th17 cells into the fetus (P < 0.0001). When pro-inflammatory Th1-or Th17-like cells were injected into syngeneic mothers, almost all of the fetuses analyzed exhibited radioactivity, suggesting that transmigration of effector T cells occurs frequently. Our results suggest the possibility of novel roles for these maternal effector cells in the pathogenesis or reduction of disease.

Painful medial knee compartment syndrome in over-45 year-olds: I--medical or surgical management: a series of 174 patients.

INTRODUCTION: There is at present no consensus on the management of degenerative medial meniscus lesions in patients aged over 45 years without proven osteoarthritis, especially given that the causal relation between degenerative meniscal lesion and osteoarthritis remains controversial. A prospective multicenter non randomized study was therefore performed. The principal objective was to assess surgeons' practice in the management of degenerative medial meniscus lesions. The secondary objectives were to identify predictive and prognostic factors and to compare medical versus surgical attitudes so as to draw up an adapted treatment strategy. PATIENTS AND METHOD: One hundred and seventy-four patients were included between September 2008 and February 2010, and distributed between a surgical (n=104) and a medical group (n=70). Minimum follow-up was 6 months. Patient satisfaction and health-related quality of life on the SF-36 questionnaire were assessed at 6 months. RESULTS: No difference emerged between the surgical and medical groups. However, predictive factors for poor results were identified: overweight (p=0.005), cartilage lesions (p=0.035) and meniscus extrusion (p=0.006). DISCUSSION: Results clarified the relation between degenerative meniscus lesions and osteoarthritis, in terms of meniscal incompetence. Meniscal extrusion should be seen as an arthrogenic degenerative meniscus lesion. We recommend a management strategy based on terrain and imaging data (X-ray and MRI), with the aim of providing patient relief while conserving cartilage.

Quality, stability, and safety data of packed red cells and plasma processed by gravity separation using a new fully integrated hollow-fibre filter device.

Background. We developed a completely closed system based on gravity separation without centrifugation steps for separation of whole blood. With this new system we compared quality and stability of the processed blood components (PRC and plasma) with respect to classical preparation. Furthermore the cost-effectiveness of this hollow fibre system was evaluated. Study Design and Methods. Whole blood collections of 15 regular blood donors were used for component preparation using the U shaped hollow fibre filter device. Results were compared to 15 whole blood preparations using centrifugation. The following parameters were evaluated: total hemoglobin, leukocyte counts, the serum concentration of total protein, lactate dehydrogenase (LDH) and potassium. Furthermore ATIII, vWF and F VIII were analyzed at different timepoints. Results. packed red cells: the data directly after separation and after 42 days of storage are in line with the guidelines of the council of Europe. Plasma. all plasma quality data are in line with the guidelines of the council of Europe for quality assurance of plasma, except for a low protein amount (factor 0.75). Conclusion. Separation of whole blood on a clinical scale in this new closed system is feasible, however the plasma protein content must be optimized.

In vitro comparison of human fibroblasts from intact and ruptured ACL for use in tissue engineering.

The present study compares fibroblasts extracted from intact and ruptured human anterior cruciate ligaments (ACL) for creation of a tissue engineered ACL-construct, made of porcine small intestinal submucosal extracellular matrix (SIS-ECM) seeded with these ACL cells. The comparison is based on histological, immunohistochemical and RT-PCR analyses. Differences were observed between cells in a ruptured ACL (rACL) and cells in an intact ACL (iACL), particularly with regard to the expression of integrin subunits and smooth muscle actin (SMA). Despite these differences in the cell source, both cell populations behaved similarly when seeded on an SIS-ECM scaffold, with similar cell morphology, connective tissue organization and composition, SMA and integrin expression. This study shows the usefulness of naturally occurring scaffolds such as SIS-ECM for the study of cell behaviour in vitro, and illustrates the possibility to use autologous cells extracted from ruptured ACL biopsies as a source for tissue engineered ACL constructs.

[Legal aspects of preparation and handling of concentrated red cells derived from placental blood in Germany]. / Juristische Aspekte bei der Herstellung und Verwendung von Plazentablut-Erythrozytenkonzentraten in Deutschland.

Legal aspects of collection, preparation and storage of autologous red cells derived from cord blood according to German law are presented and discussed.

The imbalanced mixed lymphocyte reaction between maternal and fetal lymphocytes as well as between the lymphocytes of adult children and their parents is mediated by activated CD8+ T cells.

PROBLEM: The mixed lymphocyte reaction (MLR) between maternal and fetal lymphocytes as well as between adult children and their parents is imbalanced in bidirectional mixed lymphocyte cultures. The present investigation was aimed at examining which type of T cells has primary responsibility for this phenomenon. METHODS: Two-way mixed lymphocyte cultures between lymphocytes of five female and five male adults and five male and five female newborns against lymphocytes of their parents were performed. Unrelated adults served as the controls. Before and after 72 hr incubation the mixed leukocytes cultures (MLCs) were sorted by flow cytometry according to the surface markers CD4 and CD69 or CD8 and CD69. The single cell populations within the four resulting fractions were discriminated by fluoresence in situ hybridization (FiSH) using xy-DNA probes. RESULTS: Before incubation the share of CD8(+) CD69(+) cells in the MLCs between mothers and their newborn infants was significantly shifted toward the newborn cells (P < 0.01). After 72 hr incubation the CD8(+) CD69(+) ratio in the MLCs between mother and newborn cells and between adult children (male as well female) and their parents showed a significant shift of the CD8(+) CD69(+) cells toward the children's direction. All other MLCs showed a balanced cell population for all investigated cell types, both before and after incubation. CONCLUSION: The imbalanced MLR between maternal and fetal lymphocytes and lymphocytes of adult children against their parents is mediated by a specific imbalance of the activation of CD8(+) but not of CD4(+) T cells.

Gastrointestinal symptoms and permeability in patients with juvenile idiopathic arthritis.

OBJECTIVE: Examining for gastrointestinal involvement in juvenile idiopathic arthritis is an important part of diagnostic and therapeutic procedures. Only few scientific data are available. METHODS: In a prospective study, 41 patients with juvenile idiopathic arthritis were examined for clinical and laboratory data of gastrointestinal involvement. Sugar absorption tests with lactulose, mannitol, and sucrose were applied to assess gastric and intestinal mucosal lesions. Faecal albumin and alpha 1-antitrypsin levels were measured to examine gastrointestinal protein loss, a test for occult blood in stool was administered and Helicobacter pylori serology was performed. RESULTS: 39% of our study population complained of chronic abdominal pain. The patient group showed increased sucrose excretion (p = 0.002), but a normal lactulose/mannitol ratio compared with healthy controls (p = 0.472). 21% of the patients had an elevated faecal alpha 1-antitrypsin level, but only one patient showed occult blood loss. There was no correlation between risk factors and clinical or laboratory signs of gastrointestinal involvement. CONCLUSION: We conclude that a high percentage of children and adolescents with juvenile idiopathic arthritis treated with non-steroidal antiinflammatory drugs show clinical or laboratory signs of gastrointestinal involvement.

Autologous red cells derived from cord blood: collection, preparation, storage and quality controls with optimal additive storage medium (Sag-mannitol).

To investigate whether packed red cells (PRCs) prepared from autologous cord blood-packed red cells (AC-PRCs) could be used as an alternative for homologous-packed red cells (H-PRCs), we developed a system to collect and prepare AC-PRCs and determined standard storage parameters during 35 days of storage in extended storage medium (Sag-mannitol). We collected and fractionated cord blood from 390 newborns. The amount and quality of the AC-PRCs were analysed. The bacterial contamination rate was 1.84%. Twelve AC-PRCs were stored for 35 days, and standard laboratory parameters were measured at day 1 and day 35. The initial laboratory parameters of the AC-PRCs were similar to the parameters of the H-PRCs. After 35 days, the AC-PRCs displayed an increased haemolysis rate compared to H-PRCs (1.1 versus 0.2%) and also a significant decreased adenosine triphosphate value (1.2 versus 2.3 micromol L(-1)). Haemoglobin, haematocrit and pH were comparable in both groups. AC-PRCs meet the quality criteria for H-PRCs after 35 days. Utilizing a closed collection system for cord blood and an extended storage medium will increase safety and quality and facilitate the routine transfusion of autologous red cells derived from cord blood.

Sequential changes in compliance and resistance after bolus administration or slow infusion of surfactant in preterm infants.

OBJECTIVE: As bolus instillation of surfactant can lead to acute pulmonary, hemodynamic and cerebral side effects, we tested whether pulmonary mechanics and gas exchange differ between slow surfactant infusion and bolus administration. DESIGN AND SETTING: Prospective, randomized pilot study in a tertiary care university hospital. PATIENTS AND METHODS: Of 20 consecutive preterm infants (27-35 weeks' gestation) with severe respiratory distress syndrome) who were enrolled 14 with bovine surfactant finally were analyzed. INTERVENTIONS: Six treatments were administered by slow endotracheal surfactant infusion and eight as a bolus. Static compliance (C(stat)) and resistance (R(rs)) were measured every 3 min. RESULTS: C(stat) first decreased and then increased in both groups. In the infusion group C(stat) after 90 min was significantly higher than after bolus treatment but not after 15 or 45 min. R(rs) increased about threefold, with large fluctuations in the bolus group. After 90 min PaO(2)/FIO(2) had increased from 111+/-44 to 254+/-69 in the bolus group and from 86+/-40 to 238+/-102 in the infusion group, but early FIO(2) reduction and increase in PaO(2)/FIO(2) seemed delayed in the infusion group. CONCLUSIONS: Very slow infusion of natural surfactant is at least as effective as bolus instillation in terms of improvement in C(stat) and oxygenation after 90 min. However, until 90 min the course of C(stat) and indices of gas exchange seem superior after bolus therapy. Because R(rs) is substantially increased, long expiratory times are required to yield complete exhalation.

Unknown peaks in GC-MS analyses of Guthrie cards blood samples might be the result of sample contamination by the black markings of the card.

Azelaic acid, sometimes found during analysis of Guthrie card blood samples, may be derived from the printing ink used to mark the spotting area.

Significance of respiratory syncytial virus (RSV) infection in the 1st year of life.

BACKGROUND: In this study we investigated the frequency, symptoms and predisposing factors of respiratory syncytial virus (RSV) infection during the 1st year of life in infants with obstructive airway disease in comparison with infants without airway disease. PATIENTS: We enrolled 216 infants in their 1st year of life, who were hospitalized because of obstructive airway disease. As an age- and sex-balanced control group, we examined 133 infants hospitalized for other reasons than airway disease. METHOD: A deep pharyngeal swab was taken from all infants and immediately examined for the presence of RSV antigen by using an enzyme immunoassay (Directigen). Patient data were surveyed by a questionnaire. RESULTS: The frequency of RSV infections among infants with obstructive airway disease (34.3%; n = 74) differed significantly from the control group (15%; n = 20; p < 0.01). The frequency of RSV-infected infants with obstructive airway disease decreased with age ranging from 39.1% in trimenon I to 29.0% in trimenon IV. This trend was not observed in the control group. With respect to clinical symptoms and risk factors, there were no differences between RSV-infected versus noninfected infants. CONCLUSION: RSV is an important agent causing lower obstructive airway disease (34.3% of all patients). There are no specific symptoms that can be used for diagnosing RSV infection. In order to prevent other patients on the ward from contracting nosocomial RSV infection and in the light of therapeutic options, one should test newly admitted patients presenting with symptoms of an obstructive airway disease for RSV antigen. On a ward with high-risk patients, we would recommend the use of an RSV test for all new patients.

Leukocyte adhesion deficiency II syndrome, a generalized defect in fucose metabolism.

Leukocyte adhesion deficiency II has been described in only 2 patients; herein we report extensive investigation of another patient. The physical stigmata were detected during prenatal ultrasonographic investigation. Sialyl-Lewis X (sLex) was absent from the surface of polymorphonuclear neutrophils, and cell binding to E- and P-selectin was severely impaired, causing an immunodeficiency. The elevation of peripheral neutrophil counts occurred within several days after birth. A severe hypofucosylation of glycoconjugates bearing fucose in different glycosidic links was present in all cell types investigated, demonstrating that leukocyte adhesion deficiency II is not only a disorder of leukocytes but a generalized inherited metabolic disease affecting the metabolism of fucose.

Modification of the open circuit N2 washout technique for measurement of functional residual capacity in premature infants.

We compared the standard nitrogen (N2) washout technique for measuring functional residual capacity (FRC) with a modified technique that uses a helium/oxygen mixture (heliox) at different ratio instead of pure oxygen. The tests were made with a standard lung function system equipped with an ultraviolet (UV) analyzer for measurement of N2 concentrations in the expired gas. We examined models of "spontaneous breathing" and "mechanical ventilation," each with volumes of FRC in the range of a premature and a newborn lung (20-80 ml), using both techniques at different baseline inspired oxygen concentrations (FIO2). Correlations between known and measured volumes were high and identical for the two techniques (r = 0.996), and the mean error was not significantly different from zero (P = 0.111). Measurements of FRC in 6 infants gave a correlation coefficient of r = 0.989 between the two techniques; reproducibility, as measured by the coefficient of variation, was high, showing no significant differences between both techniques (P = 0.792). However, values of individual infants were different (P = 0.011), and the slope of the regression line relating measurements by the 2 techniques was 1.04, with an intercept on the y-axis at 1.46. We conclude that FRC can be measured with the modified N2 washout technique, using heliox as a washout gas. Volumes can be measured with high precision and reproducibility, even in premature infants with low lung volumes and/or high baseline FIO2. A correction factor may be necessary to equate FRC measurements made by oxygen-N2 vs. heliox-N2 washouts. Hyperoxemia and hypoxemia can be avoided by admixing different flows of oxygen to a standard heliox mixture.

Mib-1 evaluated proliferative activity in ovarian cancer with respect to prognostic significance.

In the present study, tumor tissue of 251 patients with ovarian cancer was immunohistochemically analysed for proliferative activity using the monoclonal antibody mib-1 and the analyse system CAS 200. The rate of the growth fraction varied from 0% to 71% mib-1 positive tumor cells with a median of 17%. There was a strongly significant association between proliferative activity and the degree of histological differentiation (p = 0.0005), whereas there was no significant correlation with tumor stage and histological subtypes. Using the median as the cut-off point, patients with higher proliferating tumors (> or = 17%) had a statistically significant worse prognosis (p = 0.0431). Especially in the group of patients with grade 1 and grade 2 tumors, measurement of the proliferative activity is of help for prediction of the postoperative survival time of patients (p = 0.0336), suggesting that measurement of the growth fraction estimated by mib-1 reflects more closely the degree of tumor differentiation. However, multivariate analyses cannot confirm these results.

Yunis-Varon syndrome: the first case of German origin.

The first case of Yunis-Varon-syndrome in a child of German origin is reported. The child died at the age of 3 months. The literature is reviewed.

Intracellular acidification and Ca2+ transients in cultured rat cerebellar astrocytes evoked by glutamate agonists and noradrenaline.

The effect of different neurotransmitters on the intracellular pH (pHi) and intracellular calcium (Ca2+i) was studied in cultured astrocytes from neonatal rat cerebellum, using the fluorescent dyes 2,7'-bis(carboxyethyl)-5,6-carboxy-fluorescein (BCECF) and Fura-2. Application of glutamate or kainate (100 microM) in a HEPES-buffered, CO2/HCO3(-) -free saline induced a decrease in pHi and an increase in Ca2+i. Amplitude and time course of the pHi and Ca2+i transients were different. Glutamate and kainate evoked a mean acidification of 0.22 +/- 0.05 (n = 29) and 0.20 +/- 0.09 (n = 12) pH units, respectively. The changes in pHi and Ca2+i induced by kainate, but not by glutamate, were inhibited by 6-cyano-7-dinitroquinozalin-2,3-dion (CNQX; 50 microM). In order to elucidate the mechanism of the agonist-induced acidification, whether the pHi changes were secondary to the Ca2+ rises was tested. In the absence of extracellular Ca2+, the kainate-induced Ca2+i transient was suppressed, while the intracellular acidification was only reduced by 13%. Removal of extracellular Ca2+ reduced the glutamate-induced pHi change by 8%, while the second component of the Ca2+i transient was abolished. Application of trans-( +/- )-1-amino-(1S,3R)-cyclopentadicarboxylic acid (t-ACPD, 100 microM), a metabotropic glutamate receptor agonist, and of noradrenaline (20 microM) evoked a Ca2+i increase, but no change of pHi. D-aspartate, which has a low affinity to glutamate receptors, but is known to be transported by the glutamate uptake system in some astrocytes, evoked an intracellular acidification, similar to that induced by glutamate, but no Ca2+i transient. The results suggest that the kainate-induced acidification is only partly due to the concomitant Ca2+i rise, while the glutamate/aspartate-induced acidification is mainly due to the activation of the glutamate uptake system.

Impact on blood pressure and intestinal perfusion of dobutamine or dopamine in hypotensive preterm infants.

In a prospective study hemodynamic effects of dobutamine or dopamine (10 micrograms/kg/min) were investigated in 20 preterm infants who had protracted arterial hypotension refractory to volume therapy. Doppler ultrasonography of the superior mesenteric artery (SMA) was applied to verify intestinal perfusion and blood pressure was recorded in parallel. Mean arterial pressure (MAP) raised significantly in both groups (from 31.0 +/- 6.8 to 37.7 +/- 9.8 mm Hg during dobutamine and from 27.7 +/- 3.6 to 36.0 +/- 9.3 mm Hg during dopamine). Mean blood flow velocity increased from 25.8 +/- 13.5 to 31.5 +/- 16 cm/s with dobutamine and from 16.3 +/- 5.0 to 19.0 +/- 6.0 cm/s with dopamine (significant for dobutamine). Vascular resistance of SMA (indicated by resistance index; RI) decreased from 0.81 +/- 0.07 to 0.74 +/- 0.11 for dobutamine and from 0.89 +/- 0.06 to 0.79 +/- 0.07 for dopamine (significant for both groups). These data indicate that in the dose tested here both catecholamines are equally effective in raising MAP and lead to a significant increase of intestinal perfusion. Thus, a negative impact on mesenteric blood supply, predisposing to necrotizing enterocolitis, is not probable.

Evidence for electrogenic sodium-bicarbonate cotransport in cultured rat cerebellar astrocytes.

We have studied the regulation of intracellular pH (pHi), and HCO3(-)-dependent membrane currents in cultured astrocytes from neonatal rat cerebellum, using the fluorescent pH-sensitive dye 2,7'-bis(carboxyethyl)-5,6-carboxyfluorescein (BCECF) and the whole-cell patch-clamp technique. The steady-state pHi was 6.96 in both nominally CO2/HCO3(-)-free, HEPES-buffered saline (6.96 +/- 0.14; n = 48) and in a saline containing 5% CO2/24 mM HCO3- (6.96 +/- 0.18; n = 48) (at pH 7.4). Inhibition of the Na+/H+ exchange by amiloride (2 mM) caused a significant decrease of pHi in nominally CO2/HCO3(-)-free saline. Addition of CO2/HCO3- in the continuous presence of amiloride induced a large and fast intracellular alkalinization. Removal of external Na+ also caused a fall of pHi, and addition of CO2/HCO3- in Na(+)-free saline evoked a further fall of pHi, while the outward current was reduced or even reversed. The stilbene 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS, 0.3 mM) reduced the pHi recovery from the CO2/HCO3(-)-evoked acidification, and blocked the prominent intracellular acidification upon removal of CO2/HCO3-. Removal of external Cl- had little effect on these pHi changes. Lowering the external pH from 7.4 to 6.6 in CO2/HCO3(-)-containing saline produced a large and rapid intracellular acidification and inward current, which were both greatly reduced by DIDS and in the absence of CO2/HCO3-. The results suggest that the CO2/HCO3(-)-dependent current is partly due to a reversible bidirectional, electrogenic Na(+)-HCO3- cotransporter, which helps to regulate pHi in these cells. In addition, a prominent Na+/H+ exchanger contributes to extrude acid equivalents from these astrocytes to maintain the steady-state pHi.

Effect of recombinant human erythropoietin after allogenic bone marrow transplantation.

The hematologic effects of recombinant human erythropoietin after allogeneic bone marrow transplantation (BMT) were studied. Nineteen patients received 150 U/kg/day of C127 mouse-cell-derived recombinant human erythropoietin (rHu EPO) as a daily continuous intravenous infusion until hematocrit exceeded 35%. These data were compared with a treatment-matched historical control group of 43 patients. RHu EPO-treated patients recovered erythropoiesis more rapidly and became independent from erythrocyte transfusions after a median of 17 days, which was 7 days earlier than the control patients.