A Golgi deimpregnation study of neurons in the rhesus monkey visual cortex (Areas 17 and 18)

Summary The morhological features of 298 neurons impregnated according to Golgi-Kopsch in areas 17 and 18 of Macaca mulatta were analyzed, and the same neurons were deimpregnated to visualize structural details of the somata in different types of neurons. The following cell types were investigated: Pyramidal and pyramid-like cells, spiny stellate cells, double bouquet cells, bipolar cells, chandelier cells, neurogliaform cells, basket and related cells. This procedure allows the evaluation of the nuclear-cytoplasmic proportion and the position of the nucleus besides shape and size of the cell body. Pyramidal and pyramid-like cells (N=43), spiny stellate cells (N=26), basket and related cells (N=126) are variable in these features. A positive correlation between soma size and width of the cytoplasm is found in pyramidal, pyramid-like cells and spiny stellate cells. With the exception of some large somata in both these types of neurons the nucleus is found in a central position. Double bouquet cells (N=6), bipolar cells (N=13) and chandelier cells (N=11) exhibit small cytoplasmic rims and centrally located nuclei. The small somata of neurogliaform cells (N=37), however, and the small to very large somata of basket and related cells show broad cytoplasmic portions surrounding the eccentrically located nuclei. These findings allow the identification of different neuronal types in Nisslstained sections on the basis of these soma features. This is a prerequisite for further detailed quantitative studies on the laminar distribution of different neuronal types in the visual cortex of the monkey.     

Defective antigen presentation by Mycobacterium tuberculosis-infected monocytes.

In this study we investigated the effect of an in vitro infection with Mycobacterium tuberculosis on the ability of human monocytes to present the soluble antigen tetanus toxoid to T cells. We observed that tetanus toxoid-specific T-cell proliferation was markedly reduced when monocytes were infected with large numbers (bacterium-to-monocyte ratio, 50:1) of both viable and heat-killed mycobacteria. The level of antigen-induced gamma interferon release also was decreased when M. tuberculosis-containing monocytes were used as antigen-presenting cells. However, mycobacterium-infected monocytes did not show or trigger suppressive activity, because the presence of mycobacterium-infected monocytes did not affect the T-cell response induced by tetanus toxoid-pulsed control monocytes. When M. tuberculosis-infected monocytes were fixed with paraformaldehyde, they were not able to serve as antigen-presenting cells even in the presence of untreated accessory monocytes. Moreover, the uptake of both viable and heat-killed M. tuberculosis cells reduced the expression of human leukocyte antigen DR on monocytes. With regard to accessory function, monocytes infected with large numbers of mycobacteria were less efficient as accessory cells than were control monocytes in cultures of T cells activated with pokeweed mitogen. These results indicate that infection with large numbers of M. tuberculosis cells impairs the ability of monocytes to process and/or present soluble antigen and to serve as accessory cells in T-cell activation.     

Tumor cell growth fractions in human malignant melanomas and the correlation to histopathologic tumor grading.

The growth fraction (GF) of 72 human malignant melanomas was determined by immunostaining with monoclonal antibody Ki-67. A positive correlation of GF and histopathologically-assessed prognostic variables, such as tumor thickness, mitotic rate, and prognostic index, was found. Individual Ki-67 values were considerably scattered in all histologically defined groups of malignancy. Thus, GF as determined by Ki-67 was used to calculate a modified prognostic index. In contrast to the histologically defined prognostic index, the Ki-67 based prognostic index allows further subdivision of thin malignant melanomas with no or few mitotic figures. This may be of help in identifying tumors with a high recurrence potential.     

Abdominal pain in children and symptoms of somatization disorder

Children seen in a multispecialty medical clinic for abdominalpain were divided into three groups: 21 with confirmed organicfindings related to the abdominal pain, 14 with confirmed organicfindings unrelated to the pain, and 108 whose physical examinationswere negative (the functional pain group). For children withfunctional abdominal pain (but not for the others) the numberof symptoms of somatization disorder (Briquet's syndrome) wassignificantly related to the chronicity of the child's condition.Children with functional pain and no prior complaint had a meanof 1.95 symptoms; those with complaints of less than 1 year'sduration, 2.21 symptoms; those with complaints of more thana year since age 6, 4.04 symptoms; and those with complaintsfor more than a year with onset prior to age 6 years, 4.55 symptomsfrom the Somatization Disorder list. Findings were interpretedas preliminary evidence for a distinct, chronic, polysymptomatichysterical disorder beginning in childhood.     

Inhibitory interaction in the cat's lateral geniculate nucleus

Summary Spike activities of optic tract fibers and corresponding relay cells were recorded simultaneously in layers A and A₁ of the dorsal lateral geniculate nucleus of the cat. Light stimuli of various diameters were shone into the receptive field center of these unit pairs and their input/output ratios were determined. An increase of the stimulus size leads to an impairment of the input/output ratio in on-center and off-center relay cells. This suppressive effect has approximately the same latency as the excitatory response.Intracellular recordings suggest that the inhibitory effect of the surround is due to a postsynaptic process. Inhibitory postsynaptic potentials occur during and — under certain stimulus conditions —before the excitatory response. The short latency of these IPSPs suggests that they result from the activity of adjacent units with the same RF characteristics as the recorded neuron. This inhibitory input is not restricted to the RF periphery but may also be activated by stimulation within the RF center. Most neurons are also inhibited by units with antagonistic center responses.During the period of this research Ernst Pöppel held a training grant of the Stiftung Volkswagenwerk, Az. 11 1015.     

Production of stable anti-digoxin Fv in Escherichia coli.

We have created a bacterial expression-export system and have used it to express (14 mg l-1) the variable region fragment (Fv) of an anti-digoxin antibody (26-10) in Escherichia coli. The expression-export plasmid contains a T7 promoter and the E. coli signal sequences ompA [Movva et al., J. biol. Chem. 255, 27-29 (1980)] and phoA [Inouye et al., J. Bacteriol. 149, 434-439 (1982)] fused to heavy chain (VH) and light chain (VL) variable region sequences to generate an artificial cistron. The 26-10 Fv protein made using this system was soluble, unlike many other expression systems which produce insoluble proteins in the form of inclusion bodies. The 26-10 VH and VL proteins were cleaved at their mature N-termini and exported into the bacterial periplasm where they could be easily extracted and affinity purified on ouabain-Sepharose. 26-10 Fv bound to digoxin with similar affinity and specificity as the whole 26-10 antibody (Ka for Fv, 1.3 x 10(9) M-1, Ka for IgG, 7 x 10(9) M-1). 26-10 Fv appears to be remarkably stable in comparison with other Fv fragments. The half-life for chain dissociation of 26-10 Fv was 48 hr compared to the reported 1.5 hr half-life of McPC603 Fv. We present the proton NMR spectra of the 26-10 Fv as preliminary evidence that this expression-export system can be used to facilitate the analysis of the solution structure of 26-10 Fv by NMR.     

Differences in airway remodeling between subjects with severe and moderate asthma

BACKGROUND: Airway remodeling in asthma comprises a range of structural changes. Several studies have suggested an association between these changes and disease severity. The relationship between the extent of remodeling and lung function is not well defined. OBJECTIVE: We sought to contrast the structural changes in the airways of well-defined groups of subjects with severe and moderate asthma and to correlate the extent of remodeling with disease severity. METHODS: Endobronchial biopsy specimens were obtained from 15 subjects with severe and 13 subjects with moderate asthma. Epithelial integrity, cell-layer areas, subepithelial fibrosis, and the distance between epithelial and airway smooth muscle (ASM) layers were measured by means of image analysis. Collagen was identified by using Van Giesen stain, and ASM was defined by using smooth muscle alpha-actin immunostaining. Specific immunostains were performed for the evaluation of RANTES, IL-8, and eotaxin expression as markers of ASM phenotype. RESULTS: ASM area was greater in subjects with severe (0.24+/- 0.03 mm(2)) than in subjects with moderate (0.05+/- 0.01 mm(2)) asthma (P<.001). The distance between the epithelial and ASM layers was less in the severe group (0.12+/- 0.01 mm) than in the moderate group (0.24+/- 0.02, P<.001). A trend toward greater subepithelial fibrosis in subjects with severe asthma did not reach statistical significance. IL-8 and eotaxin expression, but not RANTES expression, were increased in the ASM of subjects with severe asthma compared with in subjects with moderate asthma. CONCLUSION: Smooth muscle alteration is the key structural change that distinguishes severe from moderate asthma, and phenotypic change in ASM might contribute to the difficulty in obtaining adequate control in some subjects with severe asthma.     

Blood Polymorphonuclear Leukocyte Activation in Atherosclerosis: Effects of Aspirin

The aim of the study was to demonstrate an activation of polymorpho-nuclear leukocytes (PMNs) in chronic progressive atherosclerosis (ATH). A group of patients with ATH, and a group of ATH patients under aspirin (ASA) therapy were compared with control persons without atherosclerotic alterations (healthy controls). Each group comprised 15 male age-matched subjects. The following inflammatory parameters related to PMN activities were measured: the polymorphonuclear leukocyte (PMN) blood count; blood PMN migration and reactive oxygen species release in vitro; the blood levels of PMN elastase, malondialdehyde, antibodies to oxidized LDL and soluble ICAM-1. In ATH patients, the PMN blood counts and the share of blood PMNs migrating upon platelet activating factor and leukotriene B4 stimulation were significnatly above the values of the healthy controls, while the other parameters were not significantly altered. ASA treatment attenuated the inflammatory response and reduced the differences between ATH and the healthy controls. It can be concluded that, in patients with chronic progressive atherosclerosis, PMNs are involved in the inflammatory process underlying the disease.     

Species-specific assessment of Mycobacterium leprae in skin biopsies by in situ hybridization and polymerase chain reaction.

Conventional histopathologic diagnosis of mycobacterial infections are limited to the determination of "acid-fast bacilli". A species-specific diagnosis is thus far impossible. In addition, routine microbiologic assessments of mycobacteria suffer from the major drawback that a species-specific diagnosis is extremely time-consuming and in several cases even impossible. As Mycobacterium leprae cannot be cultured in vitro, we tried to specifically target this obligate intracellular parasite by in situ hybridization and polymerase chain reaction (PCR) techniques. For this purpose we used a 22 mer oligonucleotide probe recognizing a species-specific sequence of the 16S rRNA of Mycobacterium leprae. Using an immunoenzymatic detection method for in situ hybridization we were able to specifically assess Mycobacterium leprae (a) in long-term cultured macrophages in vitro infected with different mycobacteria species and (b) in frozen sections of skin biopsies obtained from patients suffering from lepromatous leprosy. These results could be confirmed and extended by PCR experiments in which we used conserved oligonucleotide primers for 16S rRNA to amplify bacterial DNA isolated from different eubacterial species and from fresh-frozen as well as from formalin-fixed, paraffin-embedded and routinely processed mycobacteria-infected tissues. Upon Southern blot analysis, the Mycobacterium leprae-specific oligonucleotide probe exclusively hybridized with PCR products obtained from Mycobacterium leprae-containing samples (including paraffin sections), but not with PCR products obtained from samples containing other mycobacterial species. As species-specific oligonucleotide probes targeted at rRNA are described for a variety of mycobacterial species, these methods may be generally applied for a rapid species-specific assessment of mycobacteria in histologic material.