Umbilical cord milking in preterm infants: a systematic review and meta-analysis.

OBJECTIVE: To conduct a systematic review and meta-analysis of the efficacy and safety of umbilical cord milking in preterm infants. DESIGN: Randomised controlled trials comparing umbilical cord milking with delayed cord clamping/immediate cord clamping in preterm infants were identified by searching databases, clinical trial registries and reference list of relevant studies in November 2019. Fixed effects model was used to pool the data on various clinically relevant outcomes. MAIN OUTCOME MEASURES: Mortality and morbidities in preterm neonates. RESULTS: Nineteen studies (2014 preterm infants) were included. Five studies (n=922) compared cord milking with delayed cord clamping, whereas 14 studies (n=1092) compared milking with immediate cord clamping. Cord milking, as opposed to delayed cord clamping, significantly increased the risk of intraventricular haemorrhage (grade III or more) (risk ratio (RR): 1.95 (95% CI 1.01 to 3.76), p=0.05). When compared with immediate cord clamping, cord milking reduced the need for packed RBC transfusions (RR:0.56 (95% CI 0.43 to 0.73), p<0.001). There was limited information on long-term neurodevelopmental outcomes. The grade of evidence was moderate or low for the various outcomes analysed. CONCLUSION: Umbilical cord milking, when compared with delayed cord clamping, significantly increased the risk of severe intraventricular haemorrhage in preterm infants, especially at lower gestational ages. Cord milking, when compared with immediate cord clamping, reduced the need for packed RBC transfusions but did not improve clinical outcomes. Hence, cord milking cannot be considered as placental transfusion strategy in preterm infants based on the currently available evidence.

Efficacy of 2-Month Treatment With Cord Blood Serum Eye Drops in Ocular Surface Disease: An In Vivo Confocal Microscopy Study.

PURPOSE: To investigate the morphological changes of corneal epithelium and subbasal nerves by in vivo confocal microscopy in patients with ocular surface disease (OSD) treated with cord blood serum (CBS) eye drops. METHODS: Twenty patients with OSD (mean age 61.1 ± 12.6 years) were included in this prospective 1-arm study and treated with CBS eye drops for 2 months. Corneal sensitivity, Schirmer test score, breakup time, subjective symptoms [Ocular Surface Disease Index (OSDI) and Visual Analogue Scale (VAS)], and corneal staining were evaluated before (T0) and after (T1) treatment. In vivo confocal microscopy analyzed giant epithelial cells, subbasal nerve number and tortuosity, neuromas, beading, and dendritic cells (DCs) in the central cornea. RESULTS: OSDI, Visual Analogue Scale, and Oxford grading values significantly decreased at T1 versus T0 (respectively, 44.1 ± 18.9 vs. 74.2 ± 13.9; 3.7 ± 1.5 vs. 8.9 ± 0.9; and 2.4 ± 1.1 vs. 3.3 ± 1.3; P < 0.0001), whereas corneal sensitivity, Schirmer test score, and breakup time significantly increased (respectively, 49.5 ± 2.6 vs. 47.9 ± 2.9; 3.2 ± 2.0 vs. 2.4 ± 2.2; 4.6 ± 3.1 vs. 3.8 ± 2.1; P < 0.0001). Corneal nerve morphology improved at T1 versus T0 with a higher total nerve number (3.4 ± 1.6 vs. 2.5 ± 1.6 per frame) and lower tortuosity (3.0 ± 0.7 vs. 3.5 ± 0.6) (P < 0.01). The number of patients presenting with giant epithelial cells, beading, and neuromas decreased at T1. DC density did not change after treatment. The detection of neuromas and higher DC density at T0 were associated with greater OSDI reduction at T1 (P < 0.001). CONCLUSIONS: CBS eye drops significantly improved corneal nerve morphology and subjective symptoms in patients with severe OSD. The presence of neuromas and higher dendritic cell density at baseline were associated with greater reduction of discomfort symptoms after treatment.

Hyperbaric oxygen with cord blood transplants: Filling the donor gap.

Aljitawi OS, Paul S, Ganguly A, Lin TL, Ganguly S, Vielhauer G, Capitano ML, Cantilena A, Lipe B, Mahnken JD, Wise A, Berry A, Singh AK, Shune L, Lominska C, Abhyankar S, Allin D, Laughlin M, McGuirk JP, Broxmeyer HE. (Division of Hematologic Malignancies and Cellular Therapy; Hematology and Transplantation Translational Research Laboratory ; Pathology and Laboratory Medicine, University of Kansas Medical Center, Kansas City, Kansas; Division of Hematology/Oncology and Bone Marrow Transplantation Program, University of Rochester Medical Center, Rochester, New York; Department of Urology, University of Kansas Medical Center, Kansas City, Kansas; Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, Indiana; Cardiovascular Research Institute, Department of Biostatistics, Department of Radiation Oncology, Department of Emergency Medicine, University of Kansas Medical Center, Kansas City, Kansas; Cleveland Cord Blood Center, Cleveland, Ohio; Department of Biomedical Engineering, Case Western Reserve University, Cleveland, Ohio, USA.) Erythropoietin modulation is associated with improved homing and engraftment after umbilical cord blood transplantation. Blood 2016;128:3000-10.

Correlation of rheological parameters in maternal and fetal blood at term.

OBJECTIVE: An association between maternal and fetal blood rheology has not yet been investigated nor is it known whether and to what extent fetal blood rheology may be affected by maternal conditions. METHODS: At delivery, blood was drawn from the cubital vein of 4985 consecutive mothers and from the umbilical cord during birth for determination of blood rheological parameters (erythrocyte aggregation stasis [E0], low shear [E1], plasma viscosity [Pv]) in addition to hemoglobin (Hb) values and hematocrit (Hct). RESULTS: Maternal and newborn Pv (r = 0.2; p < 0.0001) correlated statistically significant. There was a remarkable correlation between fetal Pv and gestational age (r = 0.197; p < 0.001). Iron supplementation during pregnancy led to increased fetal Hb, Hct as well as E0 and E1 (p < 0.0001), did not have a significant impact on neonatal Pv (p = 0.068). Smoking mothers gave birth to neonates with significantly higher Pv (p = 0.049), E0 (p = 0.016) and E1 (p = 0.013). CONCLUSIONS: The increase of fetal plasma viscosity at advanced delivery time-points refers to a more gaining protein synthesis by the fetal liver and thus maturity of the fetus. Iron supplementation as well as smoking during pregnancy is associated with a relative hyper-viscosity in the fetus at delivery.

Artificial human tissues from cord and cord blood stem cells for multi-organ regenerative medicine: viable alternatives to animal in vitro toxicology.

New medicinal products and procedures must meet very strict safety criteria before being applied for use in humans. The laboratory procedures involved require the use of large numbers of animals each year. Furthermore, such investigations do not always give an accurate translation to the human setting. Here, we propose a viable alternative to animal testing, which uses novel technology featuring human cord and cord blood stem cells. With over 130 million children born each year, cord and cord blood remains the most widely available alternative to the use of animals or cadaveric human tissues for in vitro toxicology.

Diverse T-cell differentiation potentials of human fetal thymus, fetal liver, cord blood and adult bone marrow CD34 cells on lentiviral Delta-like-1-modified mouse stromal cells.

Human haematopoietic progenitor/stem cells (HPCs) differentiate into functional T cells in the thymus through a series of checkpoints. A convenient in vitro system will greatly facilitate the understanding of T-cell development and future engineering of therapeutic T cells. In this report, we established a lentiviral vector-engineered stromal cell line (LSC) expressing the key lymphopoiesis regulator Notch ligand, Delta-like 1 (DL1), as feeder cells (LSC-mDL1) supplemented with Flt3 ligand (fms-like tyrosine kinase 3, Flt3L or FL) and interleukin-7 for the development of T cells from CD34(+) HPCs. We demonstrated T-cell development from human HPCs with various origins including fetal thymus (FT), fetal liver (FL), cord blood (CB) and adult bone marrow (BM). The CD34(+) HPCs from FT, FL and adult BM expanded more than 100-fold before reaching the beta-selection and CD4/CD8 double-positive T-cell stage. The CB HPCs, on the other hand, expanded more than 1000-fold before beta-selection. Furthermore, the time required to reach beta-selection differed for the various HPCs, 7 days for FT, 14 days for FL and CB, and 35 days for adult BM. Nevertheless, all of the T cells developed in vitro were stalled at the double-positive or immature single-positive stage with the exception that some CB-derived T cells arrived at a positive selection stage. Consequently, the LSC-mDL1 culture system illustrated diverse T-cell development potentials of pre- and post-natal and adult human BM HPCs. However, further modification of this in vitro T-cell development system is necessary to attain fully functional T cells.

Characterization of osteoblastic properties of 7F2 and UMR-106 cultures after acclimation to reduced levels of fetal bovine serum.

Estrogen plays an important role in skeletal physiology by maintaining a remodeling balance between the activity of osteoblasts and osteoclasts. In an attempt to decipher the mechanism through which estrogen elicits its action on osteoblasts, experimentation necessitated the development of a culturing environment reduced in estrogenic compounds. The selected medium (OPTI-MEM) is enriched to sustain cultures under reduced fetal bovine serum (FBS) conditions and is devoid of the pH indicator phenol red, a suspected estrogenic agent. This protocol reduced the concentration of FBS supplementation to 0% through successive 24 h incubations with diminishing amounts of total FBS (1%, 0.1%, and 0%). The protocol does not appear to alter the viability, cell morphology, or osteoblast-like phenotype of 7F2 and UMR-106 cell lines when compared with control cells grown in various concentrations of FBS. Although the rate of mitotic divisions declined, the 7F2 and UMR-106 cultures continued to express osteoblast-specific markers and exhibited estrogen responsiveness. These experimental findings demonstrate that the culture protocol developed did not alter the osteoblast nature of the cell lines and provides a model system to study estrogen's antiresorptive role on skeletal turnover.

Effects of hyperbaric oxygen on proliferation and differentiation of osteoblasts from human alveolar bone.

In view of the controversy of the clinical use of hyperbaric oxygen (HBO) treatment to stimulate fracture healing and bone regeneration, we have analyzed the effects of daily exposure to HBO on the proliferation and differentiation of human osteoblasts in vitro. HBO stimulated proliferation when osteoblasts were cultured in 10% fetal calf serum (FCS), whereas an inhibitory effect of HBO was observed when cultures were supplemented with 2% FCS. On the other hand, HBO enhanced biomineralization with an increase in bone nodule formation, calcium deposition, and alkaline phosphatase activity, whereas no cytotoxic effect was detected using a lactate dehydrogenase activity assay. The data suggest that the exposure of osteoblasts to HBO enhances differentiation toward the osteogenic phenotype, providing cellular evidence of the potential application of HBO in fracture healing and bone regeneration.

Neurologic condition of healthy term infants at 18 months: positive association with venous umbilical DHA status and negative association with umbilical trans-fatty acids.

Prenatal long-chain polyunsaturated fatty acids (LCPUFAs) and trans-fatty acids may affect neurodevelopment. In healthy term children, we determined relationships between relative fatty acid contents of umbilical arteries and veins and neurodevelopment at 18 mo. The study comprised a mixed group of 317 breast-fed, formula-fed, and LCPUFA formula-fed children. Study endpoints were the Hempel neurologic examination resulting in a neurologic classification and neurologic optimality score (NOS), and the Bayley Psychomotor Developmental Index (PDI) and Mental Developmental Index (MDI). Fifteen children showed minor neurologic dysfunction (MND). The umbilical vein trans, trans-18:2n-6 content was higher in children with MND than in the normal group. The NOS was significantly reduced in infants with an umbilical vein docosahexaenoic acid (DHA) content within the lowest quartile. Umbilical vein arachidonic acid (AA) was related to NOS in univariate statistics but not in multivariate analyses. The sum of trans-fatty acids and that of C18 trans-fatty acids showed a negative association with NOS in both univariate and multivariate analyses. No associations were found between AA, DHA and total trans-fatty acids with PDI or MDI. In conclusion, neonates with a relatively low DHA status and those with high trans-fatty acid levels have a less favorable neurologic condition at 18 mo.

Autologous placental blood transfusion for the therapy of anaemic neonates.

Almost 65% of all premature neonates with a birth weight <1,500 g receive at least one erythrocyte transfusion during their first weeks of life. In the present study, we examined the feasibility of autologous transfusions in neonates, using placental blood. Placental blood was obtained from 131 of 141 preterm and term infants using a special placental blood collecting system. Approximately 20 ml of placental blood per kilogram body weight could be harvested, irrespective of birth weight. One placental blood sample was contaminated with maternal erythrocytes; aerobe or anaerobe contamination was observed in any of the stored placental blood products (n = 119) after 35 days of storage. 19 of the 141 newborns needed allogeneic erythrocyte transfusions during the first 12 weeks of life. In 5 of these 19 patients, the amount of placental blood collected would have been enough to dispense with further allogeneic blood transfusions. After completion of the preclinical study, we transfused a total of 22 children, using autologous placental blood. 8 of the 10 infants with a birth weight between 1,000 and 2,000 g and 3 of 5 infants requiring surgical intervention directly after birth needed no further allogeneic blood transfusions. We, therefore, conclude that the collection and preparation of placental blood is feasible for clinical use. The target groups of neonates who are most likely to benefit are infants with a birth weight between 1,000 and 2,000 g and neonates requiring surgical intervention directly after birth.

Sperm capacitation in vitro in the eld's deer.

Sperm capacitation was examined in the endangered Eld's deer (Cervus eldi thamin). Sperm motility and viability (percentage of sperm cells with intact membranes) were assessed in vitro over time after attempting to induce capacitation in TALP alone and TALP supplemented with calcium (10 mM CaCl2), dibutyryl cAMP (1 mM dbcAMP), or fetal calf serum (20% FCS). Sperm aliquots were evaluated at 0, 3, 6, 9, and 12 h for motility, viability, and ability to acrosome react after exposure to calcium ionophore (A23187, CI; 10 microM) or lysophosphatidylcholine (LC; 100 microg/mL). Fresh sperm aliquots in TALP + 10 mM CaCl2 exposed to CI had fewer (P < 0.05) intact acrosomes than the TALP control (TALP alone) or dbcAMP and FCS treatments after 9 h. Mean (+/- SEM) percentage of intact acrosomes of spermatozoa incubated in medium with increased CaCl2 declined (P < 0.05) from 80.2 +/- 2.6% (0 h) to 49.7 +/- 7.3% after prolonged incubation (9 h). The proportion of capacitated fresh spermatozoa was not influenced by LC treatment. Capacitation was not induced (P > 0.05) by any of the presumptive sperm capacitators after freeze-thawing. Likewise, neither CI nor LC induced the acrosome reaction (AR) in these spermatozoa, suggesting that the freeze-thawing process may have caused membrane damage. Results revealed that the supplementation of medium with CaCl2 evokes capacitation in some spermatozoa. However, Eld's deer spermatozoa appear remarkably resistant to conventional stimulators of capacitation and the AR.

Placental delivery of arachidonic and docosahexaenoic acids: implications for the lipid nutrition of preterm infants.

Arachidonic (AA) and docosahexaenoic (DHA) acids are major components of cell membranes and are of special importance to the brain and blood vessels. In utero, the placenta selectively and substantially extracts AA and DHA from the mother and enriches the fetal circulation. Studies indicate that there is little placental conversion of the parent essential fatty acids to AA and DHA. Similarly, analyses of desaturation and reductase activity have shown the placenta to be less functional than the maternal or fetal livers. There appears to be a correlation with placental size and plasma AA and DHA proportions in cord blood; therefore, placental development may be an important variable in determining nutrient transfer to the fetus and, hence, fetal growth itself. In preterm infants, both parenteral and enteral feeding methods are modeled on term breast milk. Consequently, there is a rapid decline of the plasma proportions of AA and DHA to one quarter or one third of the intrauterine amounts that would have been delivered by the placenta. Simultaneously, the proportion of linoleic acid, the precursor for AA, rises in the plasma phosphoglycerides 3-fold. An inadequate supply of AA and DHA during the period of high demand from rapid vascular and brain growth could lead to fragility, leakage, and membrane breakdown. Such breakdown would predictably be followed by peroxidation of free AA, vasoconstriction, inflammation, and ischemia with its biological sequelae. In the brain, cell death would be an extreme consequence.

17Beta-estradiol and progesterone supplementation in extremely low-birth-weight infants.

During pregnancy, 17beta-estradiol (E2) and progesterone (P) plasma concentrations increase up to 100-fold. The fetus is exposed to these increasing amounts of E2 and P. Within 1 d after delivery, E2 and P concentrations fall to nonpregnancy concentrations in the mother and the infant. Extremely premature infants are cut off from the placental supply of E2 and P at a very early developmental stage, and therefore they suffer from this deprivation for a longer period than infants born at term. Nothing is known about the consequences of this deprivation. The purpose of this study was to investigate how intrauterine concentrations of E2 and P could be maintained after birth. In 13 infants with a median gestational age of 26.4 wk (24.1-28.7), a phospholipid-stabilized soybean oil emulsion available for parenteral nutrition that contains different amounts of E2 and P was continuously administered, starting within the first postnatal hours. The supplementation was continued as long as venous access was indicated but not longer than 6 wk (median 20 d, 12-44). To maintain intrauterine plasma concentrations of 2000-6000 pg/mL E2 and 300-600 ng/mL P, 2.30 mg x kg(-1) x d(-1) E2 (1.13-3.42 mg x kg(-1) x d(-1)) and 21.20 mg x kg(-1) x d(-1) P (11.23-27.36 mg x kg(-1) x d(-1)) were needed. We conclude that supplementation of E2 and P to maintain intrauterine concentrations in extremely premature infants is possible intravenously. The infants in this study are enrolled in a randomized, controlled pilot study to evaluate the potential benefits of E2 and P supplementation.

Chlortetracycline fluorescence patterns and in vitro fertilisation of frozen-thawed boar spermatozoa incubated under various bicarbonate concentrations.

Porcine oocyte-cumulus complexes were cultured in bovine serum albumin (BSA)-free North Carolina State University (NCSU) 23 medium containing porcine follicular fluid (10%), cysteine (0.1 mg/ml) and hormonal supplements (eCG and hCG: 10 IU/ml each) for 22h. They were then cultured in the same medium but without hormonal supplements for an additional 22 h. After culture, cumulus cells were removed and oocytes were co-incubated with frozen-thawed ejaculated boar spermatozoa in tissue culture medium (TCM) 199 containing caffeine (5 mM), fetal calf serum (FCS; 10%) and varying concentrations (26-56 mM) of NaHCO3 for 9 h (experiment 1). In experiment 2, chlortetracycline (CTC) was used to assess the functional state of spermatozoa incubated under different NaHCO3 concentrations. Experiment 3 examined the effect of FCS (1% and 10%) and NaHCO3 (26 and 46 mM) on fertilisation parameters. Compared with 26 mM, penetration rate was significantly higher (p < 0.05) at 36-56 mM NaHCO3. Polyspermy showed a similar pattern although no difference was observed between 26 and 36 mM. At 46 mM NaHCO3, the mean number of spermatozoa (MNS) penetrated per oocyte increased significantly (p < 0.05). A significantly higher proportion of spermatozoa were capacitated and acrosome reacted at 46 and 56 mM NaHCO3, respectively. The fertilisation medium containing 46 mM NaHCO3 and 1% FCS showed a higher penetration rate (84%) with a relatively low incidence of polyspermy (39%). The results indicate that NaHCO3 stimulates capacitation and/or the acrosome reaction of boar spermatozoa in a dose-dependent manner and thus affects fertilisation parameters.

Corneal organ culture: effects of serum and a stabilised form of L-glutamine.

AIMS: Organ culture medium for corneas contains labile components, such as L-glutamine, whose loss could be a limiting factor to the length of storage. The medium is also supplemented with fetal bovine serum (FBS), which can vary significantly between different batches. The aim of this study was to establish the need for FBS during corneal organ culture, and to determine whether substitution of L-glutamine by the stable dipeptide L-analyl-L-glutamine was beneficial. METHODS: Porcine corneoscleral discs were suspended in 80 ml of organ culture medium (HEPES buffered Eagle's MEM with Earle's salts, 26 mmol/l NaHCO3, penicillin, streptomycin, and amphotericin B) and kept at 34 degrees C. The medium contained either 2 mmol/l L-glutamine or 2 mmol/l L-analyl-L-glutamine, and was either serum free or contained 2% FBS. At weekly intervals, five corneas from each group were stained with trypan blue and alizarin red S, and the surface area and shape of 100 endothelial cells were determined for each cornea. RESULTS: No differences were observed between corneas in organ culture medium with L-glutamine or L-analyl-L-glutamine. In serum free medium, endothelial cell density remained constant for the first week, but then declined rapidly over the next 2 weeks. With 2% FBS, there was no loss of endothelial cells for the first 2 weeks, but cell density had halved by the fourth week of organ culture. CONCLUSION: The presence of 2% FBS extended the period of endothelial stability, but no advantage was gained from the stabilised form of L-glutamine. The overall loss of endothelial cells was much greater than would be expected for human corneas.

In the absence of streptomycin, minoxidil potentiates the mitogenic effects of fetal calf serum, insulin-like growth factor 1, and platelet-derived growth factor on NIH 3T3 fibroblasts in a K+ channel-dependent fashion.

There is considerable evidence to suggest that the opening of K+ channels plays an important role in stimulating mitogenesis. K+ channel blockers have been shown to inhibit mitogenesis in vitro, mitogens increase cytosolic membrane K+ channel permeability, K+ channel openers stimulate hair growth in vivo, and the Ras/Raf signal transduction pathway induces K+ channel activity. Paradoxically, however, K+ channel openers such as minoxidil have been reported in vitro not to modulate, or even to inhibit, mitogenesis in a range of cell types. Only untherapeutic concentrations have stimulated mitogenesis. These experiments, however, appear to have been carried out in the presence of aminoglycoside antibiotics, which inhibit potassium channel activity. We now report that in the absence of aminoglycoside antibiotics, minoxidil at 10 microg/ml (0.05 mM) causes a significant stimulation of proliferation of NIH 3T3 fibroblasts maintained over a 10-d period in 5% fetal calf serum-supplemented medium. Further, we show that in the presence of 100 microg streptomycin per ml, minoxidil at 10 microg/ml produces an initial inhibition of proliferation, which apparently confirms, in NIH 3T3 fibroblasts, that the inhibition of mitogenesis by minoxidil in the presence of streptomycin is an artifact. The potentiation of NIH 3T3 cell growth by minoxidil can be attributed to the opening of potassium channels, because the potassium channel blocker tolbutamide (5 mM) or combinations of the blockers tolbutamide (1 mM)/tetraethylammonium (2 mM) or glibenclamide (1 microM)/apamin (10 nM) block the minoxidil-induced stimulation of growth. We also demonstrate that minoxidil is able to significantly potentiate the mitogenic effects of both platelet-derived growth factor and insulin-like growth factor 1 on NIH 3T3 fibroblasts in the presence of CPSR-2 (a cytokine free serum substitute). Thus we have shown that minoxidil potentiates the mitogenic effects of fetal calf serum in vitro on NIH 3T3 fibroblasts by opening potassium channels and is also able to potentiate the mitogenic effects of the growth factors platelet-derived growth factor and insulin-like growth factor 1.

Is intrapartum vibratory acoustic stimulation a valid alternative to fetal scalp pH determination?

OBJECTIVES: To determine the association between fetal heart rate accelerations, whether spontaneous or induced by vibratory acoustic stimulation, and subsequent scalp pH values in presence of a suspicious intrapartum fetal heart rate tracing, and thereby assess the ability of accelerations to predict a concurrent normal fetal scalp blood pH. DESIGN: Prospective observational study of 253 labours involving 421 pH samples. SETTING: Tertiary care university hospital of Genoeva. INTERVENTION: Vibratory acoustic stimulation through the maternal abdominal wall for five seconds prior to fetal blood sampling. MAIN OUTCOME MEASURES: Spontaneous fetal heart rate reactivity (accelerations) in the 10 min preceding vibratory acoustic stimulation, vibratory acoustic-induced reactivity prior to fetal blood sampling, and scalp pH value. RESULTS: The positive predictive value of a reactive fetal heart rate response after vibratory acoustic stimulation was 78% (95% CI 73-84%) and 97% (95% CI 94-99%) for scalp pH values of > 7.25 and > or = 7.20, respectively. Similar observations occurred with spontaneous reactivity. Of concern, 7 out of 31 (23%) occasions where the scalp blood pH was less than 7.20 appeared to be associated with a normal fetal heart rate response to vibratory acoustic stimulation. CONCLUSION: Fetal heart rate acceleration induced by vibratory acoustic stimulation was significantly associated with a normal scalp blood pH higher than 7.25. However, vibratory acoustic stimulation offers no advantage over observation of spontaneous fetal heart rate tracings and cannot safely replace fetal blood sampling during labour.

Participation of serum proteins in the inflammation-primed activation of macrophages.

Inflamed lesions release degradation products of membrane lipids, lysophospholipids, and inflamed tumor tissues release alkylglycerols. Macrophages were activated by administration of lysophosphatidylcholine (lyso-Pc) or dodecylglycerol (DDG) to mice. In vitro treatment of mouse peritoneal cells (mixture of nonadherent and adherent cells) with lyso-Pc or DDG in fetal calf serum supplemented medium for 30 min, followed by 3-h cultivation of adherent cells (macrophages) alone, resulted in greatly enhanced Fc-receptor mediated phagocytic activity and superoxide generating capacity of macrophages. The tumor lipid metabolite, DDG, is far more potent (400-fold) than lyso-Pc in terms of doses required for the maximal levels of macrophage activation. The inflammation-primed macrophage activation required a serum factor, vitamin D binding protein, as a precursor for the macrophage activating factor. Treatment of mouse peritoneal cells with 1 microgram lyso-Pc/ml or 50 ng DDG/ml in a serum-free 0.1% egg albumin supplemented medium for 30 min, followed by 3-h cultivation of the treated peritoneal cells in a medium supplemented with a very small amount (0.0005-0.05%) of ammonium sulfate [20-50% saturated (NH4)2SO4] precipitable protein fraction of FCS, resulted in greatly enhanced superoxide generating capacity of macrophages. The ammonium sulfate precipitable fraction was found to contain vitamin D binding protein.

Hyperactivated motility is coupled with interdependent modifications at axonemal and cytosolic levels in human spermatozoa.

Whether the motility characteristics of hyperactivated spermatozoa were determined by stable changes at the axonemal level and whether the presence of cytosolic factors was required for the expression of these changes was investigated. Different degrees of sperm hyperactivation were produced in Percoll-washed spermatozoa after incubation for 1 hour to 3 hours at 37 degrees C in Ham's F-10 supplemented with human blood plasma or fetal cord serum. Decomplemented fetal cord serum induced the highest percentage of hyperactivation (19 +/- 3%), followed by human plasma (13 +/- 2%). Fetal cord serum that was not decomplemented did not induce a level of hyperactivation (1.7 +/- 0.2%) significantly different from control levels (0.9 +/- 0.2%). Dialyzed fetal cord serum induced intermediate levels of hyperactivation (6 +/- 1%). The motility characteristics of demembranated sperm models of hyperactivated spermatozoa induced by decomplemented fetal cord serum and nonhyperactivated spermatozoa were compared by videomicroscopy and computer-assisted digital image analysis. After demembranation with Triton X-100 and reactivation of motility by Mg. adenosine triphosphate (Mg.ATP), hyperactivated and nonhyperactivated spermatozoa showed similar motility characteristics. However, hyperactivated spermatozoa that were demembranated and reactivated in cytosolic extracts from hyperactivated spermatozoa had significantly higher (P less than 0.05) linear velocity (33 +/- 4 mu/sec) and lower linearity (0.23 +/- 0.04) than control spermatozoa that were demembranated and reactivated in control cytosolic extracts (velocity = 24 +/- 1 mu/sec; linearity = 0.32 +/- 0.02). The data suggest that the expression of hyperactivated motility requires interdependent changes at the axonemal and cytosolic levels.(ABSTRACT TRUNCATED AT 250 WORDS)

Development potential of mouse embryos conceived in vitro and cultured in ethylenediaminetetraacetic acid with or without amino acids or serum.

Mouse ova were inseminated in vitro in modified Earle's balanced salts solution (EBSS) supplemented with 10 or 100 microM EDTA and 4 mg/ml BSA. After 4 h exposure to sperm, the ova were transferred to five different culture conditions based on albumin-free EBSS supplemented with 10 microM EDTA minus or plus amino acids, or with 100 microM EDTA minus or plus amino acids, or with human cord serum. After 44 h of culture, four-cell embryos from each culture group were transferred in cohorts of five into the left oviduct of pseudopregnant recipients (13-16 per culture condition). Two-cell embryos developed in vivo were similarly transferred to a separate group of recipients to serve as controls. The pregnancy rates following transfer of embryos cultured in 10 microM EDTA minus or plus amino acids or in 100 microM EDTA plus amino acids (38%, 43%, and 50%, respectively) were not significantly different from those of the in vivo control group (43%). The pregnancy rates following transfer of embryos cultured in 100 microM EDTA plus amino acids (21%) or plus cord serum (8%) were significantly lower (p less than 0.01) than those of the other groups. The overall yield of fetuses from total embryos transferred was significantly higher (p less than 0.01) for the groups developed in 100 microM EDTA plus amino acids (29%) and in vivo (26%) compared with embryos developed in 10 or 100 microM EDTA with no amino acids, 10 microM EDTA plus amino acids, or 100 microM EDTA plus cord serum (15%, 15%, 9%, and 3%, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)