Autonomic Nervous System Function and Essential Hypertension: Individual Response Specificity With and Without Beta-Adrenergic Blockade

The aim of the present research was to study individual response specificity in 22 male patients having essential hypertension (HT) and to compare these patients with age-matched male normotensive controls (NT). Four stimuli, letter identification, mental arithmetic, cold pressor and isometric exercise, were administered while recordings were made of: systolic and diastolic blood pressures, heart rate, respiration, forearm and hand blood flows, and skin conductance level and fluctuations. After each session urine samples were collected and epinephrine and norepinephrine levels were analyzed. Twelve subjects in the HT group were given beta-adrenergic blocking agents and retested 1 to 21 months (X?= 12 months) after the first session. Each response was standardized, using NT as the reference group. Intraclass correlations were computed to evaluate whether HT males reacted with a more consistent hierarchy of responses than did NT. Intraclass correlations were significantly higher among the patients than in the control group, regardless of whether the blood pressure response was included or excluded in the computation of the intraclass correlations. Thus, we conclude that male HT patients show more individual response specificity than NT controls. Beta-adrenergic receptor antagonists reduced levels of cardiovascular activity and attenuated reactivity but did not affect amount of specificity. Thus, intraclass correlations provide unique and useful information, since they are not related to blood pressure reactivity or to urinary catecholamine levels, nor affected by beta-adrenergic blockade.     

CX3CL1 (fractalkine) and CX3CR1 expression in myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis: kinetics and cellular origin

Background  

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS). It is associated with local activation of microglia and astroglia, infiltration of activated macrophages and T cells, active degradation of myelin and damage to axons and neurons. The proposed role for CX₃CL1 (fractalkine) in the control of microglia activation and leukocyte infiltration places this chemokine and its receptor CX₃CR1 in a potentially strategic position to control key aspects in the pathological events that are associated with development of brain lesions in MS. In this study, we examine this hypothesis by analyzing the distribution, kinetics, regulation and cellular origin of CX₃CL1 and CX₃CR1 mRNA expression in the CNS of rats with an experimentally induced MS-like disease, myelin oligodendrocyte glycoprotein (MOG)-induced autoimmune encephalomyelitis (EAE).     

A new HLA-DRB1*11 allele, DRB1*1144, identified by cloning and sequencing

We report here the identification of a novel DRB1*11 allele, DRB1*1144, identified during sequence-based HLA-DRB1 typing. Molecular cloning and direct sequencing confirmed that the new allele is identical to DRB1*110401 at exon 2, except for a single nucleotide substitution (GTG-->GCG) changing codon 38 from Valine to Alanine.     

A flattening filter for brachytherapy skin irradiation

Radioactive sources in close contact offer an alternative to superficial radiation in the treatment of skin lesions. A flattening filter was designed for a lead surface applicator to improve the skin dose distribution of a high dose rate (HDR) brachytherapy unit (Nucletron). At three heights from the opening (10, 15 and 25 mm) of the cylindrical applicator, the 192Ir source can be driven into the centre of the applicator. Thin sheets of lead foil (0.2 mm) were cut into circular shapes and placed in the opening to build a cylindrical cone that acts as a flattening filter. The shape of the cone was optimized in an iterative process using a spreadsheet and the resulting dose distribution under the applicator was determined using radiosensitive film. The use of the filter improved the dose distribution in a plane perpendicular to the beam axis to be within +/- 5% of the central axis dose. The present applicator and flattening filter together with an HDR brachytherapy unit offer an alternative for skin irradiation where a superficial unit is not available or will be replaced with a more flexible device. As the depth dose characteristics can be modified using different source-to-surface distances, the dose throughout the patient's skin can be shaped as desired by the radiation oncologist using a compensator design type approach.     

Number of interleukin-4- and interferon-γ-secreting human T cells reactive with tetanus toxoid and the mycobacterial antigen PPD or phytohemagglutinin: distinct response profiles depending on the type of antigen used for activation

The enzyme-linked immunospot (ELISPOT) assay has been proven to be an efficient and sensitive method for the enumeration of single cells secreting antibodies or cytokines. Here we have used this method to determine the number of interleukin-4 (IL-4)- and interferon-γ (IFN-γ)-producing cells in in vitro secondary responses to tetanus toxoid (TT) and the mycobacterial antigen (purified protein derivative; PPD) or the mitogen phytohemagglutinin (PHA). PHA-induced IL-4 and IFN-γ secretion was well correlated suggesting polyclonal activation of cells. This was not the case with the specific antigens, where PPD preferentially induced IFN-γ- and very few IL-4-producing cells, while TT-induced both IL-4 and IFN-γ. These differences are probably a reflection of the types of immunity the two antigens induce, mycobacteria preferentially inducing a cell-mediated T helper type 1 (Th 1) type of immunity, while immunity to tetanus is an antibody-dependent, Th 2 type of response. In individuals recently boosted with TT, a significant increase in both IL-4- and IFN-γ-producing cells in response to TT was seen at day 7 after boost, followed by decline. This was in contrast to what was seen in response to PPD where an increase of IFN-γ-producing cells after the TT boost at day 7 persisted for at least 14 days. These results suggest that after an in vivo boost both antigen-specific and nonspecific T cells are activated and that antigen-specific cells home to other organs and therefore may be difficult to demonstrate in the circulation. Our data show that the ELISPOT assay is a powerful tool for determining the frequency of cells secreting cytokines. The assay has several advantages over other assays since it is sensitive, measures the number of actually secreting cells, and avoids the problems of binding of cytokines to their cell-bound or soluble receptors.     

Looking for ferns: optimization of digestion pretreatment in fluorescence in situ hybridization (FISH) technique on paraffin-embedded tissues.

Fluorescence in situ hybridization (FISH) is a useful cytogenetic technique for the detection of chromosome aberrations. However, applying this technique routinely on paraffin-embedded tissue is hampered by technical problems. The efficiency of hybridization is influenced by formalin fixation time, and this may vary considerably between specimens. We present a simple method for improving hybridization by microscopically monitoring the time of enzymatic digestion. To establish optimal digestion time, enzymatic digestion was stopped at 3-minute intervals for biopsies and 10-minute intervals for autopsies in 24 paraffin-embedded samples. At every stop, tissue morphology was examined under light microscopy to determine if observed changes could be correlated with subsequent FISH results. The appearance of fernlike formations was found to mark the optimal digestion time that produced the strongest hybridization signals. Using this method of digestion time control, an additional 41 cases were evaluated for FISH with various types of probe. Monitoring under the microscope could be more spaced if the morphology did not change after the first visual control and could be adapted to the type of sample (in general, endoscopic samples, total digestion time of about 10 min; routine biopsies, 15 to 30 min; autopsy samples, 20 to 40 min). In every case, the appearance of the fernlike pattern correlated with proper hybridization signal. Monitoring digestion time for the appearance of fernlike structures is a useful method for improving reproducibility of FISH technique on paraffin-embedded samples. It is particularly useful when dealing with samples under heterogeneous fixation conditions (consultations, autopsies, etc.), because it eliminates the need for repetition.     

Identification of a commonly amplified 4.3 Mb region with overexpression of C8FW, but not MYC in MYC-containing double minutes in myeloid malignancies

Double minutes (dmin), the cytogenetic hallmark of genomic amplification, are found in approximately 1% of karyotypically abnormal acute myeloid leukemias (AML) and myelodysplastic syndromes (MDS). The MYC gene at 8q24 has been reported to be amplified in the majority of the cases, and generally it has been assumed that MYC is the target gene. However, only a few studies have focused on the extent of the amplicon or on the expression patterns of the amplified genes. We have studied six cases (five AML and one MDS) with MYC-containing dmin. Detailed fluorescence in situ hybridization analyses identified a common 4.3 Mb amplicon, with clustered proximal and distal breakpoints, harboring eight known genes (C8FW, NSE2, POU5FLC20, MYC, PVT1, AK093424, MGC27434 and MLZE). The corresponding region was deleted in one of the chromosome 8 homologues in five of the six cases, suggesting that the dmin originated through extra replication (or loop-formation)--excision--amplification. Northern blot analysis revealed that MYC was not overexpressed. Instead, the C8FW gene, encoding a phosphoprotein regulated by mitogenic pathways, displayed increased expression. These results exclude MYC as the target gene and indicate that overexpression of C8FW may be the functionally important consequence of 8q24 amplicons in AML and MDS.     

Human Osteogenic Sarcoma: Fine Structure of Hard Tissue Areas

The structure of hard tissue areas (with osteoid and calcified matrix) in 10 osteoblastic, chondroblastic, and fibroblastic osteogenic sarcomas was studied in the electron microscope. Neoplastic cells commonly associated with these areas and presumably actively involved in the production of hard tissue were osteo-blastlike cells types 1 and 3, chondroblastlike cells type 1, and fibroblastlike cells, as defined and characterized in previous studies. The cells differed from those in soft tissue areas of osteogenic sarcomas in but one respect: they usually showed presence of irregular extrusions at their surfaces. Other types of osteoblastlike and chondroblastlike cells occurred rarely or not at all. Two types of multinucleated giant cells were recognized in these areas, one showing a fine structure reminiscent of that in osteoclasts, the other probably being of a neoplastic nature and engaged in the production of the calcifying matrix. The evidence suggested that neoplastic osteoblastlike, chondroblastlike, and fibrolastlike cells as well as certain multincleated giant cells might all be involved in the mineralization process and/or the formation of osteoid in osteogenic sarcomas. Although phenotypically of highly variable appearance, all these different cells may thus functionally (and probably histogenetically) be closely related.

The mineralization process in the tumor tissue appeared to be a modification of what occurs in normal ossification, possibly with an alternative or complementary pathway involving the production of spherical bodies with layered contents.