Role of nuclear factor of activated T cells in chondrogenesis osteogenesis and osteochondroma formation.

PURPOSE: Nuclear factor of activated T cells (NFATc) are transcription factors that play a function in the immune response and in osteoclast differentiation. In the present work, we define the function of NFATc2 in chondrogenic and osteogenic cells. METHODS: Nfatc2loxP/loxP and Nfatc1loxP/loxP;Nfatc2loxP/loxP conditional mice were crossed with Prx1-Cre transgenics to inactivate Nfatc2 singly and with Nfatc1. Femurs and vertebrae were examined by microcomputed tomography (µCT) X-Ray images and histology and analyzed for the presence of osteochondromas. RESULTS: µCT demonstrated that Prx1-Cre;Nfatc2∆/∆ female mice had transient osteopenia and male mice did not have a cancellous or a cortical bone phenotype when compared to control mice. In contrast, the dual inactivation of Nfatc1 and Nfatc2 in Prx1-expressing cells resulted in cancellous osteopenia and small bones at 1 month of age in both sexes. Nfatc1;Nfatc2 deleted mice exhibited a ~ 50% decrease in bone volume and connectivity. Total bone area, periosteal and endocortical bone perimeters and femoral length were reduced indicating smaller bones. As the mice matured, the shortening of the femoral length persisted, but the osteopenic phenotype resolved and cancellous femoral bone of 4-month-old Nfatc1;Nfatc2 deleted mice was not different from controls although male mice had vertebral osteopenia. In addition, Nfatc1;Nfatc2 deleted mice displayed distortion of the distal metaphysis and, as they matured, the articular presence of mineralized tumors with the appearance of osteochondromas. CONCLUSION: Our studies reveal that NFATc1 and NFATc2 are necessary for optimal bone homeostasis and the suppression of osteochondroma formation.

Associations between LSAMP gene polymorphisms and major depressive disorder and panic disorder.

The purpose of this case-control genetic association study was to explore potential relationships between polymorphisms in the limbic system-associated membrane protein (LSAMP) gene and mood and anxiety disorders. A total of 21 single-nucleotide polymorphisms (SNPs) from the LSAMP gene were analyzed in 591 unrelated patients with the diagnoses of major depressive disorder (MDD) or panic disorder (PD) and in 384 healthy control subjects. The results showed a strong association between LSAMP SNPs and MDD, and a suggestive association between LSAMP SNPs and PD. This is the first evidence of a possible role of LSAMP gene in mood and anxiety disorders in humans.

Effects of bupropion augmentation on pro-inflammatory cytokines in escitalopram-resistant patients with major depressive disorder.

Studies so far have provided contradictory results on immune system markers during use of antidepressants. There are no data on changes in immune parameters after treatment augmentation. The present study aimed to clarify whether the addition of bupropion in escitalopram-resistant patients with major depression causes changes in the immune system and whether treatment response could be predicted by baseline levels of cytokines. We recruited 28 depressive patients (11 men and 17 women) who did not respond to 12-week treatment with escitalopram (20 mg/d) for an augmentation trial with bupropion (150-300 mg/day). The levels of soluble interleukin-2 receptor, interleukin-8 (IL-8) and tumor-necrosis factor-alpha were measured before and 6 weeks after addition of bupropion. For a control group, we recruited 45 healthy volunteers (19 men and 26 women). The results indicated that the baseline levels of studied cytokines did not predict treatment response to bupropion augmentation. Concentration of IL-8 increased during the treatment similarly in both responder and non-responder groups. Although bupropion augmentation had increased the response rate in escitalopram-resistant patients, this clinical improvement was not accompanied by specific changes in studied cytokine levels.

Montelukast improves asthma control in asthmatic children maintained on inhaled corticosteroids.

BACKGROUND: Because of potential toxicities of inhaled corticosteroid (ICS) use in pediatric asthma, alternative or steroid-sparing therapy is desirable. There are no previous studies evaluating montelukast's steroid-sparing effects in children with asthma. OBJECTIVE: To evaluate whether (1) montelukast as add-on therapy improves asthma symptom control and (2) montelukast provides steroid-sparing effects in children with asthma treated with low to moderate doses of ICS therapy. METHODS: In a double-blind, placebo-controlled trial, 36 children ages 6 to 14 years with symptomatic asthma maintained on a stable low to moderate dose of ICSs were randomly assigned to receive montelukast or matching placebo for 24 weeks after a run-in period of 2 weeks (period I). During the trial, subjects kept daily asthma diary cards and monthly spirometry was performed. After a 4 week add-on period (period II), the subjects completed a 20-week (period III) ICS tapering period based on a predetermined protocol. RESULTS: In period II, the difference in the number of rescue-free days was significantly higher in the montelukast group (P = 0.0001), and the number of rescue-free days per week was also significantly higher in montelukast-treated subjects compared with placebo subjects (P = 0.002). In period III, the percentage reduction in ICS dose was not significant between montelukast and placebo (P = 0.10), but the montelukast group experienced an average 17% decrease in ICS dose and the control group experienced an average 64% increase in ICS dose. CONCLUSIONS: Montelukast treatment significantly increased the number of rescue-free days in symptomatic children with asthma.

Stereotactic pallidotomy for treatment of Parkinson's disease.

There has been a resurgence of interest in surgical treatment of Parkinson's disease in the past five years due to the large number of patients who have medically intractable symptoms and because significant improvements in neuroimaging and stereotactic techniques have made surgical procedures safer and more accurate. Stereotactic pallidotomy procedures allow neurosurgeons to destroy a portion of the globus pallidus, and thereby, decrease patients' muscle rigidity from Parkinson's disease. These surgical procedures primarily involve magnetic resonance-guided stereotactic targeting and microelectrode recording techniques. To ensure successful patient outcomes, stereotactic pallidotomy procedures require special perioperative nursing interventions discussed in this article.

Immunoaffinity isolation of the sulfate conjugate of 4'-hydroxypropranolol from plasma.

Selective extraction of sulfate conjugates of basic drugs from biological matrices has been difficult because of their highly polar nature. Immunoaffinity isolation may be the best solution to this analytical problem. This was tested for a model compound, the metabolite 4'-hydroxypropranolol sulfate (HOPS), which was effectively extracted from plasma by a column containing antibodies to the parent drug propranolol. The specificity was very high, giving little interference from the biological material in subsequent high-performance liquid chromatographic analysis with fluorometric detection. The method for HOPS was highly reproducible and provided a sensitivity of 1 ng/ml plasma. The technique was applied to measurements of HOPS in plasma after therapeutic doses of propranolol as well as to the individual enantiomers after chiral derivatization.

Tumor necrosis factor alpha (TNF alpha) and transforming growth factor beta 1 (TGF beta 1) stimulate fibronectin synthesis and the transdifferentiation of fat-storing cells in the rat liver into myofibroblasts.

Transforming growth factor-beta (TGF beta 1) and tumor necrosis factor alpha (TNF alpha) stimulate the transdifferentiation of fat-storing cells (FSC) in the rat liver into highly active and "synthetic" myofibroblast-like cells (MFBIC). This activation has been documented by differential-interference contrast and light microscopy using morphologic criteria (a reduction in the number and size of fat droplets, cell flattening and the development of long cytoplasmic extensions), by the loss of retinyl-palmitate (measured by HPLC) and by the enhanced expression of iso-alpha smooth muscle actin (demonstrated by immunofluorescence microscopy). Furthermore, while cell growth measured by the cell count and DNA content is slightly inhibited by TGF beta 1 (0.81 of the control), the combination of TGF beta 1 with TNF alpha stimulates cell proliferation to 1.44 times of the control. In addition the combination of TGF beta and TNF alpha potentiated the stimulatory effect on fibronectin synthesis (TGF beta alone: 1.4 times control; TNF alpha alone: 2.2 times control; TGF beta plus TNF alpha: 4.7 times control). The total protein synthesis was not altered by TGF beta or TNF alpha. In summary the results obtained identify TGF beta and TNF alpha as mediators stimulating key events in liver fibrogenesis (i.e. FSC proliferation, FSC transdifferentiation into MFBIC, and fibronectin synthesis).

Effects of long-term treatment with lovastatin on the clotting system and blood platelets.

In a study of 20 patients with hypercholesterolemia (type IIa) the effects of lovastatin (20-80 mg/day) on various clotting and thrombosis parameters were monitored for 12 months. On 11 occasions various cholesterol fractions and clotting parameters were determined in each patient. In addition, the clotting inhibitors AT III, protein C, protein S, and C1-esterase inhibitor and the fibrinolysis parameters plasminogen and alpha 2-antiplasmin were examined. Platelet function was monitored on the basis of spontaneous and induced (collagen, ADP, epinephrine, ristocetin) aggregation. Lovastatin in the above dosage brought about a 66 mg/dl (from 320 +/- 12.6 to 254 +/- 12.0 mg/dl) reduction in the total cholesterol level and a 56 mg/dl (from 244 +/- 11.4 to 188 +/- 12.1 mg/dl) reduction in LDL cholesterol at the end of the study. Fibrinogen showed a significance decrease during the study period, whereas PT and aPTT remained unaffected. The initial slopes of the ADP-induced platelet aggregation revealed a significant decrease. C-reactive protein and platelet count remained within the normal range, indicating no significant change. Thrombin clotting time, AT III, C1-esterase inhibitor, plasminogen, and alpha 2-antiplasmin were not modified. Protein C and S behaved in a contradictory way, but remained within the normal range. Long-term treatment with lovastatin was associated with a significant reduction of fibrinogen levels and platelet aggregation induced by ADP in type-IIa hypercholesterolemic patients. These alterations, as well as their role in cardiovascular disease, should be the subject of further investigations.

Significance and quantitative analysis of von Willebrand factor in human platelets.

The von Willebrand factor (vWF) is found in plasma and in platelets. The concentration and multimeric composition of the vWF in platelets of 160 patients with bleeding tendency were examined since very little is known about the platelet vWF. For quantitative analysis of the platelet vWF, a modified ELISA was established. A reference range from 70%-130% of platelet vWF concentration considered normal was established by examining 80 healthy blood donors. 16.9% of the 160 patients showed a decreased vWF concentration in platelets only, while all the other coagulation parameters were normal. 3 of our patients belong to the same family and suggesting an autosomal dominant genetic transmission for the von Willebrand disease type 1-3. The data also suggests, that a quantitative and qualitative analysis of the vWF in plasma and platelets is required for an exact diagnosis of the von Willebrand disease.

[Diagnostic strategies for detection of the von Willebrand syndrome]. / Diagnostische Strategien zur Erkennung des von-Willebrand-Syndroms.

The von Willebrand disease (vWD) is the most severe coagulopathy. Because of the complex biochemical structure of the von Willebrand factor (vWF), a great number of types and subtypes of the vWD were found. A screening of vWD can only be done by examining the bleeding time, the ristocetin cofactor activity (risto) and by an immunological determination of the vWF concentration. Examinations of 200 patients with a bleeding tendency showed that the ratio vWF/risto < 0.7 indicates a high probability for an abnormal multimeric structure of vWF. The exact determination of the vWD subtype then has to be done by a SDS-agarose gel electrophoresis. In 16.8% of our patients we found a decreased vWF concentration in the platelets. These patients showed normal plasmatic coagulation factors, but a bleeding tendency and a prolonged bleeding time. For diagnosis of vWD the bleeding time, immunological determination of the vWF and the risto should be done first. If a ratio vWF/risto < 0.7 or a prolonged bleeding time with a bleeding tendency is found, the separation of the vWF multimers into plasma and platelets and the determination of the vWF concentration in platelets should be carried out for an exact diagnosis of vWD.

Interstitial chemotherapy with drug polymer implants for the treatment of recurrent gliomas.

Malignant gliomas have been difficult to treat with chemotherapy. The most effective agent, BCNU (carmustine), has considerable systemic toxicity and a short half-life in serum. To obviate these problems, a method has been developed for the local sustained release of chemotherapeutic agents by their incorporation into biodegradable polymers. Implantation of the drug-impregnated polymer at the tumor site allows prolonged local exposure with minimal systemic exposure. In this Phase I-II study, 21 patients with recurrent malignant glioma were treated with BCNU released interstitially by means of a polyanhydride biodegradable polymer implant. Up to eight polymer wafers were placed in the resection cavity intraoperatively, upon completion of tumor debulking. The polymer releases the therapeutic drug for approximately 3 weeks. Three increasing concentrations of BCNU were studied; the treatment was well tolerated at all three levels. There were no adverse reactions to the BCNU wafer treatment itself. The average survival period after reoperation was 65 weeks for the first dose group, 64 weeks for the second dose group, and 32 weeks for the highest dose group. The overall mean survival time was 48 weeks from reoperation and 94 weeks from the original operation. The overall median survival times were 46 weeks postimplant and 87 weeks from initial surgery. Eighteen (86%) of 21 patients lived more than 1 year from the time of their initial diagnosis and eight (38%) of 21 patients lived more than 1 year after intracranial implantation of the polymer. Frequent hematology, blood chemistry, and urinalysis tests did not reveal any systemic effect from this interstitial chemotherapy. Since the therapy is well tolerated and safe, a placebo-controlled clinical trial has been started. The trial will measure the effect of the second treatment dose on survival of patients with recurrent malignant glioma.

Reoperation for malignant astrocytoma.

We evaluated 15 consecutive patients with malignant astrocytomas who were reoperated for functional status and survival. Their Karnovsky Performance Status (KPS) was not changed by surgery. None suffered perioperative death, wound infection, or complications. Patients with glioblastoma maintained KPS unchanged for a mean of 13 weeks (median, 10 weeks); with anaplastic astrocytoma, mean, 37.2 weeks (median, 24 weeks). Life spans were approximately twice that of non-reoperated historical controls. Reoperation for patients with recurrent malignant astrocytoma should be seriously considered when a gross total re-resection can be the goal in a patient whose tumor is in an accessible brain region.

Quantitative analysis of 6-keto-prostaglandin F1 alpha using immunoaffinity purification and gas chromatography-mass spectrometry.

This paper describes an immunoaffinity purification technique for 6-keto-prostaglandin F1 alpha (6KPGF1 alpha) prior to quantitative analysis by high-resolution gas chromatography-negative-ion chemical ionization mass spectrometry (HRGC-NICIMS). Polyclonal antibodies to 6KPGF1 alpha were partially purified using Staphylococcus aureus Protein A immobilized on Sepharose CL-4B. This partially purified fraction was covalently bound to silica gel using N-hydroxysuccinimidyl-functionalized silica. Columns constructed using this gel quantitatively bound 6KPGF1 alpha which could be eluted quantitatively with acetonitrile-water (19:1). Binding capacity was reconstituted by washing with 0.01 M phosphate buffer (pH 7.4). Human urinary and canine plasma 6KPGF1 alpha was sufficiently purified using these columns that HRGC-NICIMS analysis of the methoxime-pentafluorobenzyl-tris-trimethylsilyl derivative was interference-free.

Revascularization for moyamoya disease: five-year follow-up.

We present a case of severe moyamoya disease in a 4-year-old child involving the anterior and posterior circulations. Encephaloduroarteriosynangiosis was performed and the child was followed for 5 years with serial angiography. Early clinical stabilization was attained and the child has normal or superior intellectual development despite her early fixed deficits. Follow-up angiography revealed the development of several large direct anastomotic channels.

Tethered cervical spinal cord. Case report.

A case of congenital tethered cervical spinal cord is presented in a young adult. Metrizamide computerized tomography was the most useful imaging technique for identifying the tethered spinal cord. Intraoperative somatosensory evoked potentials correlated well with clinical improvement following surgery.

Spontaneous reduction of a closed depressed skull fracture: a case report.

Presented is a single case of a 6-year-old child in whom a closed depressed skull fracture was found to be elevated spontaneously less than 2 days after the injury. There were no cosmetic or neurological sequelae at follow-up. This case demonstrates that these injuries do not always require operation and that they occasionally resolve spontaneously, even in school-age children.

Extraction of thromboxane B2 from urine using an immobilized antibody column for subsequent analysis by gas chromatography-mass spectrometry.

This paper describes an antibody affinity (immunoaffinity) column which, in one step, extracts and sufficiently purifies urinary thromboxane B2 (TXB2) for quantitative analysis by high resolution gas chromatography-negative ion chemical ionization-selected ion monitoring-mass spectrometry (HRGC-NICI-SIM-MS). Polyclonal TXB2 antibody from rabbit was partially purified using immobilized Staphylococcus aureus Protein A. The purified IgG fraction was then immobilized using an N-hydroxysuccimidyl silica gel. The resulting matrix bound 570 ng TXB2 per ml of gel. TXB2 was quantitatively eluted with acetonitrile-water (19:1). Columns constructed from the gel could be used repeatedly since binding capacity was reconstituted using 0.01 M phosphate buffer (pH 7.4) with no apparent loss of activity. Using these columns, urinary TXB2 was sufficiently purified in one step such that in subsequent analysis by HRGC-NICI-SIM-MS interference free chromatograms were observed.