Effects of IFN-gamma and interleukin-1beta on major histocompatibility complex antigen and intercellular adhesion molecule-1 expression by 9L gliosarcoma: relevance to its cytolysis by alloreactive cytotoxic T lymphocytes.

To enhance the efficacy of cellular immunotherapy for gliomas, we tested the concept of using proinflammatory cytokine treatment with interferon-gamma (IFN-gamma) or interleukin-1beta (IL-1beta) or both to render glioma cells more susceptible to cytolysis by alloreactive cytotoxic T lymphocytes (aCTL). The cytokines, separately or in combination, were able to upregulate major histocompatibility complex (MHC) class I antigen or intercellular adhesion molecule-1 (ICAM-1) on Fischer rat 9L gliosarcoma cells. 9L cells were incubated in vitro for 24, 48, or 72 h with varying concentrations of rat IFN-gamma (0-2000 U/ml) or recombinant human IL-1 (rHUIL-1) (0-1000 U/ml) or both. By 48 h, IFN-gamma (500 U/ml) maximally induced the percentage of positive expressing cells and the relative antigen density of MHC class I and ICAM-1 on 9L cells, whereas IL-1 induced only ICAM-1 expression. Simultaneous incubation of IL-1 with IFN-gamma did not further affect the induction of class I on 9L cells more than that achieved with IFN-gamma alone. 9L cells with upregulated MHC class I and ICAM-1 expression were more sensitive to lysis by aCTL in in vitro cytotoxicity assays, regardless of whether the precursor aCTL came from naive or from 9L-immunized rats. Furthermore, inhibition of 9L cytotoxicity in assays that included blocking antibodies to MHC class I or to ICAM-1 revealed that T cell receptor (TCR) interactions with MHC class I and that ICAM-1 interactions with lymphocyte function-associated-1 (LFA-1) antigen account for a portion of the glioma lysis by aCTL.     

Corpectomy for multi-level cervical spondylosis and ossification of the posterior longitudinal ligament

The choice of a surgical approach for multi-level cervical spondylotic myelopathy (CSM) and ossification of the posterior longitudinal ligament (OPLL) is still a controversial issue. While most of the surgeons are still performing decompression by laminectomy some are doing multi-level anterior decompression. Few neurosurgeons are performing decompression by corpectomy. We have treated 26 patients by median cervical corpectomy during the last 4 years. These patients were followed up for a mean period of 25 months. Twenty one (80%) patients had a good outcome, 2 patients remained unchanged and 3 expired. Review of the literature and our experience indicates that patients with CSM and OPLL should be operated by median cervical corpectomy (anterior approach).     

Pancreatoblastoma (infantile pancreatic carcinoma)--a case report

Pancreatoblastoma or infantile pancreatic carcinoma is a rare pancreatic tumor with distinct acinar and squamoid cell differentiation that generally affects infants and young children. Ultrasound and CT scan may be useful but preoperative diagnosis is often quite difficult. The outcome is generally favourable. A such case of 10 years old boy with an abdominal mass is being presented.     

Intrathoracic pressure fluctuations move blood during CPR: Comparison of hemodynamic data with predictions from a mathematical model

Whether blood flow during cardiopulmonary resuscitation (CPR) results from intrathoracic pressure fluctuations or direct cardiac compression remains controversial. We developed a mathematical model that predicts that blood flow due to intrathoracic pressure fluctuations should be insensitive to compression rate over a wide range but dependent on the applied force and compression duration. If direct compression of the heart plays a major role, however, the model predicts that flow should be dependent on compression rate and force, but above a threshold, insensitive to compression duration. These differences in hemodynamics produced by changes in rate and duration form a basis for determining whether blood flow during CPR results from intrathoracic pressure fluctuations or from direct cardiac compression. The model was validated for direct cardiac compression by studying the hemodynamics of cyclic cardiac deformation following thoracotomy in four anesthetized, 21–32-kg dogs. As predicted by the model, there was no change in myocardial or cerebral perfusion pressures when the duration of compression was increased from 15% to 45% of the cycle at a constant rate of 60/min. There was, however, a significant increase in perfusion pressures when rate was increased from 60 to 150/min at a constant duration of 45%. The model was validated for intrathoracic pressure changes by studying the hemodynamics produced by a thoracic vest (vest CPR) in eight dogs. The vest contained a bladder that was inflated and deflated. Vest CPR changed intrathoracic pressure without direct cardiac compression, since sternal displacement was <0.8 cm. As predicted by the model and opposite to direct cardiac compression, there was no change in perfusion pressures when the rate was increased from 60 to 150/min at a constant duration of 45% of the cycle. Manual CPR was then studied in eight dogs. There was no surgical manipulation of the chest. Myocardial and cerebral blood flows were determined with radioactive microspheres and behaved as predicted from the model of intrathoracic pressure, not direct cardiac compression. At nearly constant peak sternal force (378–426 N), flow was significantly increased when the duration of compression was increased from short (13%–19% of the cycle) to long (40%–47%), at a rate of 60/min. Flow was unchanged, however, for an increase in rate from 60 to 150/min at constant compression duration. In addition, myocardial and cerebral flow correlated with their respective perfusion pressures. Thus vital organ perfusion pressures and flow for manual external chest compression are dependent on the duration of compression, but not on rates of compression of 60 and 150/min. These data are of course similar to those produced by vest CPR, where intrathoracic pressure is manipulated without sternal displacement, and to those predicted for movement of blood by intrathoracic pressure changes. These data are, however, opposite to those produced by cardiac deformation and to those predicted for movement blood by direct cardiac compression. We conclude that intrathoracic pressure fluctuations generate blood flow during manual CPR.     

Levetiracetam Inhibits both ryanodine and IP3 receptor activated calcium induced calcium release in hippocampal neurons in culture

Epilepsy affects approximately 1% of the population worldwide, and there is a pressing need to develop new anti-epileptic drugs (AEDs) and understand their mechanisms of action. Levetiracetam (LEV) is a novel AED and despite its increasingly widespread clinical use, its mechanism of action is as yet undetermined. Intracellular calcium ([Ca²⁺]_i) regulation by both inositol 1,4,5-triphosphate receptors (IP3R) and ryanodine receptors (RyR) has been implicated in epileptogenesis and the maintenance of epilepsy. To this end, we investigated the effect of LEV on RyR and IP3R activated calcium-induced calcium release (CICR) in hippocampal neuronal cultures. RyR-mediated CICR was stimulated using the well-characterized RyR activator, caffeine. Caffeine (10 mM) caused a significant increase in [Ca²⁺]_i in hippocampal neurons. Treatment with LEV (33 μM) prior to stimulation of RyR-mediated CICR by caffeine led to a 61% decrease in the caffeine induced peak height of [Ca²⁺]_i when compared to the control. Bradykinin stimulates IP3R-activated CICR—to test the effect of LEV on IP3R-mediated CICR, bradykinin (1 μM) was used to stimulate cells pre-treated with LEV (100 μM). The data showed that LEV caused a 74% decrease in IP3R-mediated CICR compared to the control. In previous studies we have shown that altered Ca²⁺ homeostatic mechanisms play a role in seizure activity and the development of spontaneous recurrent epileptiform discharges (SREDs). Elevations in [Ca²⁺]_i mediated by CICR systems have been associated with neurotoxicity, changes in neuronal plasticity, and the development of AE. Thus, the ability of LEV to modulate the two major CICR systems demonstrates an important molecular effect of this agent on a major second messenger system in neurons.     

Enhanced molecular volume of conservatively pegylated Hb: (SP-PEG5K)6-HbA is non-hypertensive

Recent studies have suggested that the "pressor effect" of acellular Hb is a consequence of perturbation of the macro-and microcirculatory system in multiple ways, and that PEGylation is an effective approach for controlling the same. In an attempt to confirm this concept, a new and simple thiolation mediated, maleimide chemistry-based conservative PEGylation protocol has been developed to conjugate multiple copies of PEG-chains to Hb. This approach combines the high reactivity of maleimides towards thiols with the propensity of iminothiolane to derivatize the epsilon-amino groups of proteins into reactive thiol groups, with conservation of their positive charge. One of the PEGylated products, namely (SP-PEG5K)6-HbA, that carries on an average six copies of PEG5000 chains per Hb, is non-hypertensive in hamster top load and in rat 50% exchange transfusion models. This hexa-PEGylated-Hb has (i) a hydrodynamic volume corresponding to that of an oligomerized Hb of 256kDa, (ii) a molecular radius of approximately 6.8 nm, (iii) high oxygen affinity, (iv) lowered Bohr effect, and (v) increased viscosity and colloidal osmotic pressure. These properties of (SP-PEG5K)6-HbA are consistent with the emerging new paradigms for the design of Hb based oxygen carriers and confirm the concept that the "pressor effect" of Hb is a multifactorial event. The thiolation mediated maleimide chemistry-based PEGylation protocol described here for the generation of (SP-PEG5K)6-Hb is simple, highly efficient, and is carried out under oxy conditions. The results demonstrate that a non-hypertensive PEG-Hb can be generated by conjugation of a lower number of PEG chains than previously reported.     

Altered chromatin landscape in circulating T follicular helper and regulatory cells following grass pollen subcutaneous and sublingual immunotherapy

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Primary malignant lymphoma of the breast

Primary malignant lymphomas of the breast (PBL) are uncommon constituting only 0.04 to 0.53% of malignant breast neoplasms. We wish to report the clinical, cytological and histologic findings of PBL diagnosed in a 52 years old female.     

Human MSH2 binds to trinucleotide repeat DNA structures associated with neurodegenerative diseases

The expansion of trinucleotide repeat sequences is associated with several neurodegenerative diseases. The mechanism of this expansion is unknown but may involve slipped-strand structures where adjacent rather than perfect complementary sequences of a trinucleotide repeat become paired. Here, we have studied the interaction of the human mismatch repair protein MSH2 with slipped-strand structures formed from a triplet repeat sequence in order to address the possible role of MSH2 in trinucleotide expansion. Genomic clones of the myotonic dystrophy locus containing disease-relevant lengths of (CTG)n x (CAG)n triplet repeats were examined. We have constructed two types of slipped-strand structures by annealing complementary strands of DNA containing: (i) equal numbers of trinucleotide repeats (homoduplex slipped structures or S-DNA) or (ii) different numbers of repeats (heteroduplex slipped intermediates or SI-DNA). SI-DNAs having an excess of either CTG or CAG repeats were structurally distinct and could be separated electrophoretically and studied individually. Using a band-shift assay, the MSH2 was shown to bind to both S-DNA and SI-DNA in a structure- specific manner. The affinity of MSH2 increased with the length of the repeat sequence. Furthermore, MSH2 bound preferentially to looped-out CAG repeat sequences, implicating a strand asymmetry in MSH2 recognition. Our results are consistent with the idea that MSH2 may participate in trinucleotide repeat expansion via its role in repair and/or recombination.      

Regulatory Role of Vitamin D Receptor Gene Variants of BsmI, ApaI, TaqI, and FokI Polymorphisms on Macrophage Phagocytosis and Lymphoproliferative Response to Mycobacterium tuberculosis Antigen in Pulmonary Tuberculosis

The regulatory role of vitamin D receptor (VDR) gene variants of Bsm I, Apa I, Taq I, and Fok I polymorphisms on vitamin D(3)-modulated macrophage phagocytosis with live Mycobacterium tuberculosis and lymphoproliferative response to M. tuberculosis culture filtrate antigen (CFA) was studied in patients with pulmonary tuberculosis (n = 46) and in normal healthy subjects (NHS) (n = 64). Vitamin D(3) at a concentration of 1 x 10(-7) M enhanced the phagocytic potential of normal subjects who had a phagocytic index of less than 20%. This increase was seen in subjects with the genotypes BB (p = 0.017), AA (p = 0.016), tt (p = 0.034), and FF (p = 0.013) and the extended genotype BBAAtt (p = 0.034). Normal subjects with BBAAtt performed better phagocytosis than individuals with bbaaTT genotype (p = 0.034). Vitamin D(3) at 10(-9), 10(-8), and 10(-7) M concentrations suppressed the lymphoproliferative response to CFA antigen in normal subjects. This decreased lymphocyte response was observed in normal individuals with the genotypes BB (p = 0.0009), tt (p = 0.016), and FF (p = 0.008) and the extended genotype BBAAtt (p = 0.02). Addition of vitamin D(3) had no significant effect on macrophage phagocytosis and lymphoproliferative response to CFA in pulmonary TB patients. This may be due to the unresponsive nature of the cells to the action of vitamin D(3) or the downregulated VDR expression by virtue of the disease, which renders them inactive. The genotypes BB, tt, and the extended genotype BBAAtt may be associated with increased expression of VDR which in turn regulate the action of vitamin D(3) and modulate the immune functions to M. tuberculosis in NHS.