Spontaneous generation of functional osteoclasts from synovial fluid mononuclear cells as a model of inflammatory osteoclastogenesis

In osteoimmunology, osteoclastogenesis is understood in the context of the immune system. Today, the in vitro model for osteoclastogenesis necessitates the addition of recombinant human receptor activator of nuclear factor kappa‐B ligand (RANKL) and macrophage colony‐stimulating factor (M‐CSF). The peripheral joints of patients with rheumatoid arthritis (RA) and spondyloarthritis (SpA) are characterized by an immune‐mediated inflammation that can lead to bone destruction. Here, we evaluate spontaneous in vitro osteoclastogenesis in cultures of synovial fluid mononuclear cells (SFMCs) activated only in vivo. SFMCs were isolated and cultured for 21 days at 0.5–1.0 × 10⁶ cells/mL in culture medium. SFMCs and healthy control peripheral blood monocytes were cultured with RANKL and M‐CSF as controls. Tartrate‐resistant acid phosphatase (TRAP) positive multinucleated cells were found in the SFMC cultures after 21 days. These cells expressed the osteoclast genes calcitonin receptor, cathepsin K, and integrin β3, formed lacunae on dentin plates and secreted matrix metalloproteinase 9 (MMP9) and TRAP. Adding RANKL and M‐CSF potentiated this secretion. In conclusion, we show that SFMCs from inflamed peripheral joints can spontaneously develop into functionally active osteoclasts ex vivo. Our study provides a simple in vitro model for studying inflammatory osteoclastogenesis.     

Heterogeneity of cerebral vasoreactivity in preterm infants supported by mechanical ventilation

The reaction of cerebral blood flow to acute changes in arterial carbon dioxide pressure (PaCO2) and mean arterial blood pressure was determined in 57 preterm infants supported by mechanical ventilation (mean gestational age 30.1 weeks) during the first 48 hours of life. All infants had normal brain sonograms at the time of the investigation. In each infant, global cerebral blood flow was determined by xenon-133 clearance two to five times within a few hours at different levels of PaCO2. Changes in PaCO2 followed adjustments of the ventilator settings. Arterial oxygen pressure was intended to be kept constant, and mean arterial blood pressure fluctuated spontaneously between measurements. The data were analyzed by stepwise multiple regression, with changes in global cerebral blood flow, PaCO2, mean arterial blood pressure, and postnatal age or intracranial hemorrhage used as variables. In infants with persistently normal brain sonograms, the global cerebral blood flow-carbon dioxide reactivity was markedly lower during the first day of life (mean 11.2% to 11.8%/kPa PaCO2) compared with the second day of life (mean 32.6/kPa PaCO2), and pressure-flow autoregulation was preserved. Similarly, global cerebral blood flow-carbon dioxide reactivity and pressure-flow autoregulation were present in infants in whom mild intracranial hemorrhage developed after the study. In contrast, global cerebral blood flow reactivity to changes in PaCO2 and mean arterial blood pressure was absent in infants in whom ultrasonographic signs of severe intracranial hemorrhage subsequently developed. These infants also had about 20% lower global cerebral blood flow before hemorrhage, in comparison with infants whose sonograms were normal, a finding that suggests functional disturbances of cerebral blood flow regulation. Several perinatal factors were tested, but only birth after abruptio placentae was related to subsequent periventricular hemorrhage (p = 0.037).     

Early growth delay in diabetic pregnancy: relation to psychomotor development at age 4

Ninety nine consecutive insulin dependent and 101 non-diabetic pregnant women were examined by ultrasonograph to assess early fetal growth. In 42 of the diabetic mothers and three of the non-diabetic mothers the scan showed early intrauterine growth delay. At 4-5 years of age all children available for study were evaluated by the Denver developmental screening test. Only 23 of the 34 children of diabetic mothers with early intrauterine growth delay had normal test scores compared with 46 of the 50 children of diabetic mothers with normal intrauterine growth. The children failed in personal-social development, gross motor development, and particularly in language and speech development. Children of diabetic mothers with normal early fetal growth had scores very similar to those of the children of non-diabetic mothers, of whom 76 of the 86 tested had normal scores. This study suggests that children with a history of growth delay in early diabetic pregnancy should be screened for possible developmental impairment.     

The lack of effects of zinc and nitric oxide in initial state of pilocarpine-induced seizures.

In this study we investigated whether intracerebroventricular (i.c.v.) injection of L-NAME (a nitric oxide synthase inhibitor) or CaEDTA (an extracellular zinc chelator) or the combination of the two could affect the initial phase of pilocarpine induced (2 h) seizures. Two groups of rats were used. Animals from both groups were given with i.c.v. injections of either saline (10 microl), L-NAME (150 microg/10 microl), CaEDTA (100 mM/10 microl) or L-NAME and CaEDTA. One group received pilocarpine HCl (380 mg/kg i.p.) the other served as control. Pilocarpine HCl was injected intraperitoneally 10 min later. The behavior of the animals was observed for 2h and the intensity of their seizures was scored. The rats were then sacrificed and their brains were removed and analyzed for zinc ions by using the immersion autometallography and the TSQ fluorescence staining. All the animals which received pilocarpine HCl developed seizures. Despite treatment with L-NAME and/or CaEDTA we found that the latency and the intensity of seizures were similar in both groups investigated. The distribution of stainable zinc ions and the intensity of staining in hippocampus were not affected by pilocarpine and found unchanged after L-NAME and/or CaEDTA injections in both the control animals and the pilocarpine treated animals. The data suggest that the nitric oxide system and zinc ions do not affect pilocarpine-induced seizures in their initial state.     

Expression of Human Recombination Activating Genes (RAG-1 and RAG-2) in Lymphoma

Two recently discovered genes, the recombination activating genes 1 and 2 (RAG-1 and RAG-2), are necessary to perform variable (V), diversity (D), and joining (J) recombination. They synergistically activate VDJ recombination to generate immunocompetent lymphocytes. Disruption of either gene results in a maturation arrest at a very early B and T cell progenitor stage. Expression and downregulation of RAG's are closely associated with interleukin 7, sIgM and TCR-CD3 complex, respectively. Assessment of RAG mRNA expression is a valuable marker in identifying the genotypic maturation status of leukemias and lymphomas. Persistent RAG expression in otherwise mature lymphoid proliferations may explain puzzling biological and clinical observations such as multiple rearrangements in lymphomas with a mature phenotype. Lack of RAG expression in Hodgkin's disease with abundant Reed-Stern-berg cells is consistent with a mature phenotype of the latter. Availability of a anti-RAG-1 monoclonal antibody in the near future will facilitate RAG analysis of lymphomas.     

Immunocytochemical localization of zinc transporter 3 in the ependyma of the mouse spinal cord

We report, for the first time, the light microscopical and ultrastructural appearance of ZnT3-immunoreactivities in the ependymal cells of the central canal of the mouse spinal cord. Light microscopy revealed the presence of ZnT3-immunoreactive (Ir) ependymal cells in 1 microm thick epon sections stained by the ABC method. The ZnT3-Ir cells were observed at all levels of the spinal cord, but were a little more numerous in lumbosacral segments than in cervicothoracic segments. The ZnT3-Ir cells had large, ovoid nuclei with abundant cytoplasm, and protruded into the lumen of the central canal. Our ultrastructural findings suggest that the ZnT3-Ir ependymal cells possess secretory activity directed towards the central canal. We propose that they may play a role in the trans-ependymal mechanism responsible for zinc homeostasis between cerebrospinal fluid and the central area of the gray matter.     

Effects of androstenedione, insulin and luteinizing hormone on steroidogenesis in human granulosa luteal cells

BACKGROUND: The present study was conducted to investigate the effect of androstenedione, insulin and LH on human granulosa cell oestrogen and progesterone production. We postulated that elevated concentrations of androstenedione, insulin and LH may be important modulators of granulosa cell steroidogenesis. METHODS: Granulosa cells obtained in connection with IVF procedures were cultured for a total of 4 days in serum-free medium containing androstenedione (10(-6) mol/l). We tested the effect of androstenedione (10(-5) mol/l) on insulin (0-800 microIU/ml), LH (1-10 ng/ml) as well as on insulin + LH-stimulated oestrogen and progesterone production. RESULTS: Insulin increased the basal secretion of steroid hormones, and furthermore augmented LH-stimulated oestrogen and progesterone accumulation in granulosa cell cultures. Androstenedione (10(-5) mol/l) stimulated basal oestrogen production, but significantly reduced (32-58%) insulin + LH-stimulated oestrogen and progesterone secretion (P < 0.05). CONCLUSION: These results suggest that high androstenedione concentrations may act directly to impair insulin augmentation of LH-stimulated oestradiol and progesterone production in cultured human granulosa luteal cells. This is compatible with the hypothesis that high androgen levels may inhibit oestrogen production in polycystic ovary follicles, and as such may contribute to anovulation and infertility in women with polycystic ovary syndrome.     

Ultrastructural localization of gold in macrophages and mast cells exposed to aurothioglucose

The ultrastructural localization of gold in resting, stimulated, and activated mouse peritoneal cells exposed in vivo and in vitro to aurothioglucose was examined by a photochemical technique. Activated macrophages from herpes simplex virus type 2 infected mice showed the heaviest accumulation of gold, located in phagolysosomes. Gold was also visualized in granules of mast cells and in nuclei of disintegrated cells. No gold was found in polymorphonuclear leucoctyes and in lymphocytes. The mechanism by which gold is taken up into the cells is discussed.     

Neonatal outcome in a Danish national cohort of 3438 IVF/ICSI and 10,362 non-IVF/ICSI twins born between 1995 and 2000

BACKGROUND: In Denmark, one-third of twin pregnancies are the result of IVF/ICSI treatment. Limited data on neonatal outcome in IVF/ICSI twins are available in the literature. METHODS: A register study was conducted on neonatal morbidity and mortality in a complete national twin cohort including all 3438 (3393 live-born) IVF/ICSI and 10,362 (10,239 live-born) non-IVF/ICSI twins born between 1995 and 2000. Twins were identified in the National Medical Birth Registry and dichotomized into IVF/ICSI and non-IVF/ICSI by cross-reference with the Danish IVF Registry. Data on neonatal morbidity and mortality were retrieved from the Danish Patient Registry and the Danish Registry of Causes of Deaths. In order to exclude monozygotic twins, sub-analyses on unlike-sex twins were conducted. RESULTS: A birth weight discordance of >20% was observed in 20.6% of IVF/ICSI versus 15.7% of control twin pairs (P < 0.001). The risk of discordant birth weight >20% was OR 1.29 (95% CI 1.04-1.58) in unlike-sex IVF/ICSI twins versus control twins. The risk of delivery at <37 completed weeks and birth weight <2500 g was similar in the two cohorts; however, in unlike-sex IVF/ICSI versus control twins the risk of delivery at <37 weeks and birth weight <2500 g was OR 1.22 (95% CI 1.09-1.38) and OR 1.25 (1.11-1.40) respectively. After stratification for maternal age and parity, these risks disappeared. IVF/ICSI twins carried a higher risk of admittance to a neonatal intensive care unit (NICU) than control twins (OR 1.18, 95% CI 1.09-1.27), and this was even more pronounced in unlike-sex twins [OR 1.34 (95% CI 1.19-1.51)]. No differences were observed in malformation or mortality rates between the two cohorts. CONCLUSIONS: Despite higher birth weight discordance and more NICU admissions among IVF/ICSI twins, neonatal outcome in IVF/ICSI twins seems to be comparable with that of non-IVF/ICSI twins, when only dizygotic twins were considered in the comparisons.