Microheterogeneity of acute phase proteins in patients with clinically active and clinically nonactive osteoarthritis

Summary Microheterogeneity of two acute phase glycoproteins, -1-acid glycoprotein (AGP) and -1-antichymotryspin (ACT), concentrations of AGP, ACT, and C-reactive protein (CRP), and levels of three cytokines: interleukin 1 (IL-1-), interleukin 6 (IL-6), and tumor necrosis factor (TNF-) were determined in 61 serum samples and 7 synovial fluids (SFs) obtained from patients (n=61) with osteoarthritis. Using affinity immunoelectrophoresis with concanavalin A (conA), a significant decrease in the reactivity of AGP and ACT with this lectin was found in patients with clinically active osteoarthritis when compared to those with clinically nonactive disease (p<0.001 and p<0.05, respectively). There was no increase in the concentration of AGP, ACT, and C-reactive protein (CRP) in the sera examined. In particular, no increase in the serum level of these proteins was found in the patients with clinically active disease. Low concentrations of IL-6 and TNF- were found in most sera and SFs examined. In 6 out of 7 SFs available, IL-6 concentrations were higher than in the respective serum samples but for TNF- the same could be shown in one case only. Low concentrations of IL-1- were found in 4 serum samples obtained from patients with clinically active osteoarthritis and in no SF specimen studied. In the entire group, serum level of TNF- correlated weakly with the AGP and ACT reactivity coefficients with conA (r=0.3634, p<0.005 and r=0.3324, p<0.02, respectively).Our findings suggest that there are changes in the microheterogeneity of acute phase glycoproteins in some patients with osteoarthritis similar to those observed in rheumatoid arthritis and other chronic inflammations. Possible mechanisms of the involvement of cytokines in the regulation of glycosylation of acute phase glycoproteins in osteoarthritis are discussed.     

Tumor-associated lymphocytes (TAL) are competent to produce higher levels of cytokines in neoplastic pleural and peritoneal effusions than those found in sera and are able to release into culture higher levels of IL-2 and IL-6 than those released by PBMC

This work was designed to study the proliferative response of tumor-associated lymphocytes (TAL) from neoplastic effusions against autologous tumor cells and the immunophenotype pattern of TAL from neoplastic effusions and that of PBMC of the same patients. We also compared the serum levels of the cytokines interleukin (IL) 1, 2 and 6, tumor necrosis factor- (TNF) and soluble IL-2 receptor (sIL-2R) with those present in neoplastic effusions of the same patients. Moreover, we examined the ability of TAL and peripheral blood mononuclear cells (PBMC) to produce and release the cytokines and sIL-2R and to express membrane CD25 following their stimulation with phytohemagglutinin (PHA) in vitro. Finally, we compared the cytokines/sIL-2R production and membrane CD25 expression by PHA-stimulated PBMC of the patients with neoplastic effusions with a series of 90 cancer patients without neoplastic effusions and 20 normal healthy subjects. Thirteen neoplastic pleural and eight peritoneal effusions were collected from 11 patients with primary lung cancer, 7 with primary epithelial ovarian cancer, 1 with breast cancer, 1 with pleural mesothelioma, and 1 with pancreatic cancer. The proliferative response of TAL from neoplastic effusions against autologous tumor cells was lower than the response to PHA, IL-2, and anti-CD3, but significant. The percentage distribution of CD3⁺ and CD8⁺ lymphocyte subpopulations was higher in peritoneal than in pleural effusions, while the CD16⁺ subset was higher in pleural than in peritoneal effusions. The percentage distribution of CD16⁺ was significantly lower in pleural effusions than in PBMC of patients with pleural effusions. The CD39 antigen was higher on TAL from peritoneal effusions than on PBMC of the same patients. The levels of IL-1 and sIL-2R in peritoneal effusions did not differ from those measured in the sera of the same patients, while the levels of IL-2, IL-6, and TNF were higher in the peritoneal effusions. The levels of IL-2, IL-6, TNF, and sIL-2R, but not IL-1, in pleural effusions were significantly higher than those found in the sera of the same patients. The amounts of IL-2 and IL-6 produced by TAL were generally higher than those released by PBMC. The secretion of cytokines IL-1, IL-2, and sIL2R by PHA-stimulated PBMC was lower, but IL-1 and IL-6 secretion was higher in cancer patients with neoplastic effusions than in either cancer patients without neoplastic effusions or normal subjects. The CD25 expression on PHA-stimulated PBMC derived from cancer patients with neoplastic effusions was in the same range as that of cancer patients without neoplastic effusions and normal subjects. These findings suggest that TAL may be able to produce cytokines and may be amenable to immune manipulation.Abbreviations FITC Fluorescein-isothiocyanate - IL Interleukin - mAb Monoclonal antibody - MHC Major histocompatibility complex - NK Natural killer - PBMC Peripheral blood mononuclear cells - PHA Phytohemagglutinin - TAL Tumor-associated lymphocytes - TIL Tumor-infiltrating lymphocytes - TNF Tumor necrosis factor- - sIL-2R Soluble interleukin-2 receptor     

The selective P2X purinoceptor agonist, β,γ-methylene-L-adenosine 5′-triphosphate,discriminates between smooth muscle and neuronal P2X purinoceptors

The effects of the putative selective P_2X purinoceptor agonist, ,-methylene-l-adenosine 5-triphosphate (me-l-ATP), were determined at rat neuronal and smooth muscle P_2X purinoceptors.Me-l-ATP had no effect on the extracellularly recorded membrane potential of the rat isolated vagus nerve preparation at concentrations up to 300 M. In contrast, the archetypal P_2X purinoceptor agonist, , methylene ATP (meATP;1–100 M), produced concentration-related depolarisation responses with a mean EC₅₀ value of 10.8 M. The depolarising effects of meATP were not attenuated by me-l-ATP (100 M). In voltage clamp experiments on single nodose ganglion neurones, ATP (100 M), but not me-l.-ATP (1–300 M), evoked rapid ( < 20 ms onset) inward currents when applied using a concentration-clamp method. In receptor binding studies to rat brain membranes, me-d-ATP and meATP competed with high affinity for [³H]Lx meATP binding sites, with mean pIC₅₀ values of 7.7 and 8.3, respectively. However, me-l-ATP possessed low affinity for these sites and competed only at concentrations in excess of 10 M (mean pIC₅₀ value 4.1).In prostatic segments of the rat vas deferens, me-l-ATP (1–100 M) and meATP (0.3–100 M) each produced concentration-related contractile responses with mean EC₅₀ values of 17.1 and 3.6 M, respectively. Me-l-ATP (1–10 M) evoked fast inward currents in freshly dispersed vas deferens smooth muscle cells, indicative of an action at ligand-gated ion channels. Binding sites in vas deferens membranes labelled using 1 nM [³H]meATP exhibited high affinity for me-l-ATP, meATP and me-d-ATP with mean PIC₅₀ values of 7.7, 8.4 and 7.3, respectively.These results indicate that me-l-ATP exhibits neither agonist nor antagonist properties at P_2X purinoceptors on rat vagal neurones and possesses only very low affinity for [³H]meATP binding sites in rat brain. In contrast, me-l-ATP is a potent, high affinity agonist at smooth muscle P_2X purinoceptors of the rat vas deferens. This selective agonist action of me-l-ATP suggests that P_2X purinoceptors in smooth muscle and neurones are different and represent distinct P_2X purinoceptor subtypes.     

Transient decrease in sound tolerance levels following hearing deprivation in normal-hearing subjects

ObjectiveTo determine the circadian influence on sound sensitivity produced by temporal hearing deprivation in healthy normal human subjects.DesignParticipants underwent bilateral earplugging before completion of anthropometry, the author's developed questionnaire, the Hamilton Anxiety and Depression Inventory, pure tone audiometry (PTA), stapedial reflex thresholds (SRT), distortion products otoacoustic emissions input/output (DPOAE-I/O), and uncomfortable loudness levels (ULLs). Afterward, the participants were randomly divided into group A, starting at 8:00 a.m. and finishing at 8:00 p.m., and group B, starting at 4:00 p.m. and ending at 4:00 a.m. Serum cortisol levels and audiological test results were obtained at the beginning and end of the session and 24-h free urinary cortisol levels were measured.Study sampleThirty healthy volunteers.ResultsPTA was 2.68 and 3.33 dB HL in groups A and B, respectively, with no statistical difference between them. ULLs were significantly lower in group A compared to group B, with an average of 8.1 dB SPL in group A and 3.3 dB SPL in group B (p < 0.0001). A SRT shift was observed in group A, with no difference in group B, and a night shift in DPOAE-I/O in group B.ConclusionsReduced loudness tolerance is demonstrated during daytime hearing deprivation in contrast to nighttime; this may be due to increased central gain in the awake cortex.     

Auswirkungen von Argon-Gasembolien während eines PneumoperitoneumsRID="*"ID="*"Die Studie wurde zum Teil gefördert von der DFG (Bo 1329/8–1)

Zusammenfassung. In einer experimentellen Studie wurde bei 10 Schweinen mit einem mittleren K?rpergewicht von 18,9 (15–24) kg eine intraven?se CO₂- oder Argon-Embolie mit 10, 20 und 30 ml Gas durchgeführt. Das invasive Monitoring zeigte bei der Gasembolie mit Argon im Gegensatz zur Gasembolie mit CO₂ einen st?rkeren Anstieg des pulmonal arteriellen Drucks (p < 0,001), einen st?rkeren Abfall des endexspiratorischen CO₂ (p < 0,01), des Herzminutenvolumens (p < 0,01) und des mittleren arteriellen Drucks (p < 0,01). In der Argon-Gruppe (n = 5) starben zwei Tiere nach 20 bzw. 30 ml Bolusgabe. Ein weiteres Tier konnte nach Gabe von 30 ml Bolus erfolgreich reanimiert werden. In der CO₂-Gruppe (n = 5) starb weder eines der Tiere noch war eine Reanimation erforderlich. Wenig l?sliche Gase wie Argon sollten in Situationen mit erh?htem Risiko einer Gasembolie nicht angewendet werden. ID=" Dr. T. Junghans Klinik f&uuml;r Allgemein-, Visceral-, Gef&auml;&szlig;- und Thoraxchirurgie Universit&auml;tsklinikum Medizinische Fakult&auml;t der Humboldt-Universit&auml;t Campus Charit&eacute; Mitte Schumannstra&szlig;e 20/21 D-10117 Berlin     

Specific inhibition of epidermal growth factor receptor tyrosine kinase by 4-anilinoquinazolines

Summary Since the mitogenic action of EGF is mediated by ligand-induced autophosphorylation of the EGF receptor (EGFR), and EGFR is commonly overexpressed in solid human tumours, inhibitors of receptor tyrosine kinase activity (RTK) could prove to be effective antitumour agents. Screening of a compound library using an EGF-RTK enzyme prepared from human tumour derived A431 cells identified a series of potent (IC₅₀<1µM) enzyme inhibitors. These inhibitors are quinazolines bearing a variety of substituted anilines at the 4-position. The most potent 4-anilinoquinazolines (IC₅₀ 20nM) have small non-polar meta substituents on the aniline ring, and are competitive with ATP and non-competitive with substrate. The growth inhibitory activity of these agents was assessed in vitro using KB cells (human oral squamous tumour) grown in the absence or presence of EGF. A selected compound, 4-(3-chloroanilino)quinazoline (CAQ), inhibited EGF-stimulated growth in a concentration dependent manner and complete blockade was observed at concentrations (1–10 µM) which had no effect on basal growth. Selectivity of growth inhibition by CAQ was further exemplified in IGF1-stimulated KB cells where no effect was detected at concentrations which completely blocked EGF-stimulated growth. Similarly, CAQ blocked TGF-stimulated growth in MCF-7 human breast cancer cells without affecting insulin-stimulated growth. These studies define a novel class of EGF-RTK inhibitors which are also potent and selective inhibitors of EGF-stimulated human tumour cell growthin vitro. Presented at the symposium "New Approaches in the Therapy of Breast Cancer", Georgetown University Medical Center, Washington DC, October 1994, generously supported by an education grant from Bristol-Myers Squibb.     

Interleukin-1β effects on cyclic GMP and cyclic AMP in cultured rat islets of Langerhans — arginine — dependence and relationship to insulin secretion

Summary When islets were cultured with interleukin-1 (1 or 100 pmol/l) for 12 h in arginine-containing medium, cyclic GMP levels were increased 1.6- and 4.5-fold respectively. The arginine analogue, N--nitro-l-arginine methyl ester, which blocks nitric oxide formation and partially reverses inhibition of insulin secretion by 100 pmol/l interleukin-1, largely, but not completely, blocked generation of cyclic GMP. Treatment of islets with 100 pmol/l interleukin-1 for 12 h significantly decreased islet cyclic AMP generation in the absence of isobutylmethylxanthine (from 13.1±0.7 to 9.3±0.8 fmol/g islet protein), this fall was arginine-dependent and may have resulted from an effect on a cyclic AMP phosphodiesterase, since it was masked if isobutylmethylxanthine was present. Isobutylmethylxanthine (0.4 mmol/l) reduced the inhibitory potency of interleukin-1 in 15 h slightly but significantly from 80.5 to 59.0%. The morpholinosydnonimine SIN-1, which is a nitric oxide donor, inhibited insulin secretion, raised islet cyclic GMP and lowered cyclic AMP; its effects were similar to those of interleukin-1. However, 6-anilinoquinoline-5,8-quinone, [LY83583 (1–10 mol/l)], inhibited insulin secretion, and significantly decreased cyclic GMP while 8-bromocyclic GMP stimulated insulin secretion. Both low- and high-dose interleukin-1 treatment give a large arginine-dependent and a small, yet significant, arginine-independent increase in cyclic GMP. The inhibitory effect of SIN-1 or interleukin-1 on insulin secretion seems to depend to a small extent on decreased islet cyclic AMP, though sustained increases in nitric oxide or depleted islet GTP may directly affect the secretory process.     

α-Smooth-muscle actin expression in normal and fibrotic human livers

To determine the significance of the expression of -smooth-muscle actin in the fibrotic human liver, normal and diseased livers were stained with anti--smooth-muscle-actin antibody by an immunoperoxidase method. Vitamin A-containing lipocytes were also identified by the modified Kupffer's gold chloride method. In the normal human liver, lipocytes as well as vascular smooth muscle cells expressed -smooth-muscle actin. In alcoholic liver disease, there was an increase in the cells positive for -smooth-muscle actin adjacent to the fibrotic areas, but the response of lipocytes to the gold chloride reaction diminished. In chronic hepatitis, the cells positive for -smooth-muscle actin increased around the enlarged portal areas, and the response to the gold chloride reaction did not change appreciably. An increase in the cells positive for -smooth-muscle actin was associated with the progression of hepatic fibrosis in the liver of patients with alcoholic liver disease and chronic hepatitis.     

Role of brainstem and spinal noradrenergic and adrenergic neurons in the development and maintenance of hypertension in spontaneously hypertensive rats

Summary In young and adult spontaneously hypertensive rats (SHR), dopamine -hydroxylase (DBH) and phenylethanolamine N-methyltransferase (PNMT) activities in discrete areas of the brainstem and spinal cord were measured as indices of noradrenergic and adrenergic neuronal activities. In young SHR, the DBH activities were elevated in the locus coeruleus (LC), A2 cell area and thoracic intermediolateral cell area (IML). The elevation disappeared at adult SHR. In young SHR, no significant change of PNMT activity was observed in the A1, A2, nucleus tractus solitarii (NTS), LC and IML areas, while, in adult SHR, the PNMT activity in the A1 cell area and DBH activity in the NTS were elevated. Lowering of blood pressure by hydralazine decreased the PNMT activity elevated in the A1 cell area and elevated it in the NTS.Plasma levels of norepinephrine and epinephrine, as measured in blood samples collected via aortic cannula at resting state, were much lower than many reported values in blood collected from the decapitated trunk. In young SHR, a significant elevation of plasma norepinephrine and DBH levels was confirmed as signs of peripheral sympathetic nervous activation. The elevation disappeared at adult SHR. Plasma epinephrine levels raised under restraint stress were much higher in SHR at all ages than in normotensive controls.In young SHR, the selective activation of noradrenergic neurons of the IML, A2 and LC areas, accompanied by activation of the peripheral sympathetic nervous system, initiates the hypertension. In adult SHR, the activation of adrenergic neurons in the A1 cell area including the nucleus reticularis lateralis may not be involved in the maintenance of hypertension but may be the results of hypertension.