LAT (linker for activation of T cells): a useful marker for megakaryocyte evaluation on bone marrow biopsies

Detection of atypical megakaryocytes in bone marrow biopsies, especially in cases of myelodysplastic syndromes (MDS), chronic myeloproliferative disorders (CMPD) and acute leukemias, is facilitated by staining for markers such as Ulex europaeus agglutinin (UEA)-J, CD31, CD61 and von Willebrand factor (VWF), the latter being considered the most sensitive. Recently, LAT (linker for activation of T cells), a molecule involved in T-cell activation and platelet aggregation, was found to be expressed by megakaryocytes and platelets in tissue sections. We compared VWF and LAT immunoreactivity on megakaryocytes in 64 bone marrow biopsies from 12 normal controls (NC), and from patients with MDS (n=18), CMPD (n=21) and acute megakaryocytic leukemia (AML-M7, n=13). Immunostaining was performed on paraffin sections with polyclonal antibodies against VWF and LAT. Immunoreactivity was evaluated by counting positive megakaryocytes in 10 high-power fields, and values were compared using Student's t test for paired data. Both VWF and LAT predominantly stained the cytoplasm of megakaryocytes, although LAT was also recognizable on the cell membrane. In most biopsies, the immunoreactivity of the two antibodies was quite similar. No significant differences were noticed between the mean values of VWF+ and LAT+ megakaryocytes. However, in 22 cases (5 NC; 5 MDS; 6 CMPD; 6 AML-M7), the number of LAT+ megakaryocytes was at least 30% higher than VWF+cells, while in 3 cases opposite findings were found. In 3 AML-M7 cases, anti-LAT antibodies stained numerous megakaryocytes, but anti-VWF staining was practically negative; in another 5 AML-M7 cases, anti-LAT labeling was much stronger than anti-VWF staining. LAT represents a useful immunohistochemical marker for megakaryocytes in normal and pathological conditions. It seems to be expressed by megakaryocytes more than VWF in most cases and, particularly, in conditions associated with poorly differentiated megakaryocytes, such as acute megakaryocytic leukemias. The use of LAT staining should be recommended in association with other megakaryocyte markers in the study of bone marrow biopsies in cases of hematopoietic disorders.     

Substance P (neurokinin-1) and neurokinin A (neurokinin-2) receptor gene and protein expression in the healthy and inflamed human intestine

Increasing evidence suggests that tachykinins are involved in the control of pathophysiological states, such as inflammation. The precise localization of tachykinin receptors is of paramount importance in the search for their possible physiological and pathological role; in this study, therefore, we attempted to define cellular sites of substance P (NK-1R) and neurokinin A (NK-2R) receptor expression in the healthy and the inflamed human intestine by in situ hybridization and immunohistochemistry. In the normal ileum and colon, NK-1R and NK-2R were localized to smooth muscle cells of the muscularis mucosae and propria and a few inflammatory cells of the lamina propria; NK-1R expression was also found in the muscular wall of submucosal blood vessels, enteric neurons and, to a lesser degree, in surface epithelial cells. Patients with Crohn's disease and ulcerative colitis showed a dramatic increase in NK-1R density relative to controls, in both the inflamed and the uninvolved mucosa. Up-regulation of NK-1R was particularly evident on epithelial cells lining the mucosal surface and crypts, as well as on endothelial cells of capillaries and venules. Also, a marked increase in NK-2R expression was found in both groups of patients on inflammatory cells of the lamina propria, especially eosinophils. Our findings demonstrate that in the normal human intestine NK-1R and NK-2R are expressed in multiple cell types, which are endowed with different physiological functions; in addition, they demonstrate that both NK-1R and NK-2R are up-regulated in patients with Crohn's disease and ulcerative colitis. Taken together, these observations may have important physiological and pathophysiological implications, and provide the rationale for the use of NK-1R and NK-2R antagonists in the treatment of inflammatory bowel disease.     

Expression of platelet-derived growth factor and its receptors in normal human liver and during active hepatic fibrogenesis.

Expression of platelet-derived growth factor (PDGF) and its receptor (R) subunits was evaluated in normal human liver and in cirrhotic liver tissue by in situ hybridization and immunohistochemistry. In normal liver, PDGF and PDGF-R subunit expression was limited to a few mesenchymal cells of the portal tract stroma and vessels. In cirrhotic liver, PDGF-A and -B chain mRNA expression was markedly increased and was co-distributed with immunoreactivity for PDGF-AA and -BB in infiltrating inflammatory cells and along vascular structures within fibrous septa. These aspects were paralleled by a marked overexpression of PDGF-R alpha- and beta-subunit mRNAs and of the relative immunoreactivities in a wide range of mesenchymal cells in fibrous septa and in perisinusoidal alpha-smooth-muscle-actin-positive cells. In general expression and distribution of PDGF-R subunits appeared to be related to the activation of different mesenchymal cell types involved in the fibroproliferative process. Therefore, we evaluated the expression of PDGF-R subunits in liver tissue specimens with increasing degrees of necroinflammatory activity. The results of this additional study confirmed that expression of PDGF-R subunits is highly correlated with the severity of histological lesions and collagen deposition. Our results, providing evidence for a functional involvement of PDGF/PDGF-R in liver fibrogenesis, greatly support the results of previous in vitro studies and direct attention toward pharmacological strategies able to affect the series of signaling events arising from the autophosphorylation of PDGF-R subunits.     

A role for the basal ganglia in nicotinic modulation of the blink reflex

Summary In humans and rats we found that nicotine transiently modifies the blink reflex. For blinks elicited by stimulation of the supraorbital branch of the trigeminal nerve, nicotine decreased the magnitude of the orbicularis oculi electromyogram (OOemg) and increased the latency of only the long-latency (R2) component. For blinks elicited by electrical stimulation of the cornea, nicotine decreased the magnitude and increased the latency of the single component of OOemg response. Since nicotine modified only one component of the supraorbitally elicited blink reflex, nicotine must act primarily on the central nervous system rather than at the muscle. The effects of nicotine could be caused by direct action on lower brainstem interneurons or indirectly by modulating descending systems impinging on blink interneurons. Since precollicular decerebration eliminated nicotine's effects on the blink reflex, nicotine must act through descending systems. Three lines of evidence suggest that nicotine affects the blink reflex through the basal ganglia by causing dopamine release in the striatum. First, stimulation of the substantia nigra mimicked the effects of nicotine on the blink reflex. Second, haloperidol, a dopamine (D₂) receptor antagonist, blocked the effect of nicotine on the blink reflex. Third, apomorphine, a D₂ receptor agonist, mimicked the effects of nicotine on the blink reflex.     

Synergy between the genes for butyrylcholinesterase K variant and apolipoprotein E4 in late-onset confirmed Alzheimer's disease

The allelic frequency of the gene for the K variant of butyrylcholinesterase (BCHE-K) was 0.17 in 74 subjects with late-onset (age > 65 years) histopathologically diagnosed Alzheimer's disease (AD), which was higher than the frequencies in 104 elderly control subjects (0.09), in 14 early-onset cases of confirmed AD (0.07) and in 29 confirmed cases of other dementia (0.10). The association of BCHE-K with late-onset AD was limited to carriers of the epsilon 4 allele of the apolipoprotein E gene (APOE), among whom the presence of BCHE-K gave an odds ratio of confirmed late-onset AD of 6.9 (95% C.I. 1.65-29) in subjects > 65 years and of 12.8 (1.9-86) in subjects > 75 years. In APOE epsilon 4 carriers over 75 years, only 1/22 controls, compared with 10/24 confirmed late-onset AD cases, had BCHE-K. We suggest that BCHE-K, or a nearby gene on chromosome 3, acts in synergy with APOE epsilon 4 as a susceptibility gene for late-onset AD.      

Dosimetric verification of a commercial 3D treatment planning system for conformal radiotherapy with a dynamic multileaf collimator

The dosimetric accuracy of a 3D treatment planning system (TPS) for conformal radiotherapy with a computer-assisted dynamic multileaf collimator (DMLC) was evaluated. The DMLC and the TPS have been developed for clinical applications where dynamic fields not greater than 10 x 10 cm2 and multiple non-coplanar arcs are required. Dosimetric verifications were performed by simulating conformal treatments of irregularly shaped targets using several arcs of irradiation with 6 MV x-rays and a spherical-shaped, tissue-simulating phantom. The accuracy of the delivered dose at the isocentre was verified using an ionization chamber placed in the centre of the phantom. Isodose distributions in the axial and sagittal planes passing through the centre of the phantom were measured using double-layer radiochromic films. Measured dose at the isocentre as well as isodose distributions were compared to those calculated by the TPS. The maximum percentage difference between measured and prescribed dose was less than 2.5% for all the simulated treatment plans. The mean (+/-SD) displacement between measured and calculated isodoses was, in the axial planes, 1.0 +/- 0.6 mm, 1.2 +/- 0.7 mm and 1.5 +/- 1.1 mm for 80%, 50% and 20% isodose curves, respectively, whereas in the sagittal planes it was 2.0 +/- 1.2 mm and 2.2 +/- 2 mm for 80% and 50% isodose curves, respectively. The results indicate that the accuracy of the 3D treatment planning system used with the DMLC is reasonably acceptable in clinical applications which require treatments with several non-coplanar arcs and small dynamic fields.     

The distribution of neuropeptide Y-immunoreactive neurons and nerve fibers in the forebrain of the carp Cyprinus carpio L

The present study reports the distribution of Neuropeptide Y (NPY)-immunoreactive neurons and fibers in the forebrain of the adult carp Cyprinus carpio L. Serial Nissl-stained sections were used for cytoarchitecture and identification of anatomical structures. Immunostaining of NPY-containing neurons and fibers was used as neurochemical marker and tool for comparison with other species, including the goldfish.The general outline of the cytoarchitecture of the carp forebrain is similar to that of other Cypriniformes. However, using NPY immunohistochemistry, we found several specific differences with the goldfish, especially in the diencephalon. In the hypothalamus of the carp NPY-immunoreactive (NPYir) neurons were identified in the n. dorsolateralis thalami, and in the n. ventralis lateralis thalami. In the same location, we observed the n. anterior hypothalami and the n. preglomerulosus pars lateralis, described in the goldfish, as parts of n. prerotundus. However, in the carp we were not able to identify a n. preglomerulosus pars medialis, a n. preglomerulosus pars medialis commissuralis and a n. glomerulosus. We describe a n. rotundus, in which we did not find substructures typical of the goldfish.Further differences with the goldfish, trout and salmon were also noted.     

The salivary microbiota as a diagnostic indicator of oral cancer: A descriptive,non-randomized study of cancer-free and oral squamous cell carcinoma subjects

Background  

The purpose of the present investigation was to determine if the salivary counts of 40 common oral bacteria in subjects with an oral squamous cell carcinoma (OSCC) lesion would differ from those found in cancer-free (OSCC-free) controls.