Laboratory strategies for efficient handling of paraffin-embedded tissues for molecular detection of clonality in non-hodgkin lymphomas.

We herein present a technical strategy to optimize DNA isolation from paraffin-embedded tissue (PET). This includes the choice of adequate buffers for proteinase K digestion and multiplex PCR amplifications for assessing the appropriateness of DNA extracts for subsequent PCR assays for detecting clonality. We found that the association of proteinase K digestion in nonionic buffer and subsequent extract dilutions accounted for 79% of successful amplifications. A final efficiency of 88% was achieved by additional organic extractions and/or re-extractions. Comparisons were carried out with control DNA extracts from fresh samples to assess the efficiency of each clonality assay. Immunoglobulin CDRIII rearranged region amplification was more efficient for pregerminal center B-cell lymphomas in contrast to CDRII rearrangement detection, which was more effective for germinal and postgerminal lymphomas. T-cell clonality detection by TCRgamma PCR was less efficient in PET samples than in fresh tissues showing that DNA integrity is more critical for TCR than for IGH amplification. Two inconclusive cases without phenotypic markers and two other atypical lymphoproliferations masked by reactive T cells were diagnosed as plasmablastic lymphomas and as monoclonal B-proliferations, respectively, due to IGH rearrangements.     

Immunomodulation of autoimmune and inflammatory diseases with intravenous immunoglobulin

Abstract Intravenous immunoglobulin (IVIg) has been used in the treatment of primary and secondary antibody deficiencies for over two decades. Since the early 1980s, the therapeutic efficacy of IVIg has been established in idiopathic thrombocytopenic purpura, Guillain-Barré syndrome, chronic inflammatory demyelinating polyneuropathy, myasthenia gravis, dermatomyositis and Kawasaki syndrome, and the prevention of graft versus host disease in recipients of allogeneic bone marrow transplants. Its use has also been reported in a large number of other autoimmune and systemic inflammatory conditions. In this review, we discuss the mechanisms by which IVIg exerts immunomodulatory effects in immune pathologies.     

Defining epithelial cell progenitors in the human oxyntic mucosa

In the human stomach, the oxyntic epithelium includes numerous tubular invaginations consisting of short pits opening into long glands. The pit is lined by pit cells, whereas the gland is composed of three regions: the base, containing zymogenic cells; the neck, containing neck cells; and the isthmus, composed of little known immature cells and of parietal cells, which are also scattered through the neck and base. The aim of this study was to examine the ultrastructure of the immature cells and to determine their relation to mature cells. To do so, normal oxyntic mucosal biopsies from subjects ranging from 20-43 years old were fixed in aldehydes and postfixed in reduced osmium for electron microscopy and morphometric analysis. The immature cells were sorted out into four classes, whose roles were clarified by comparison with the thoroughly investigated mouse oxyntic epithelium. The first class was composed of the least differentiated immature cells, which were rare and characterized by minute, dense, or cored secretory granules and were accordingly named mini-granule cells. Their function was not clarified. The second class consisted of pre-pit cells, which were characterized by few dense mucous granules and give rise to pit cells that ascend the pit wall and, after reaching the luminal surface, die or are extruded. Both pre-pit and pit cells underwent continuous renewal and, therefore, together constituted a renewal system referred to as pit cell lineage. The third class, or pre-neck cells, characterized by cored secretory granules, give rise to neck cells that descend toward the base region and differentiate further into pre-zymogenic cells, which finally become zymogenic cells. The latter eventually degenerate and die. Thus pre-neck cells and their progeny constitute a renewing system, designated zymogenic cell lineage. The fourth class, or pre-parietal cells, characterized by long microvilli and few tubulovesicles, differentiate into parietal cells that descend along the neck and base regions and eventually degenerate and die. Pre-parietal and parietal cells represent a renewing system referred to as parietal cell lineage. While the origin of the last three classes of progenitor cells has not been elucidated, it is likely that they arise either from an unidentified multipotential stem cell, possibly the mini-granule cell itself, or from the mitotic activity of pre-pit and pre-neck cells. In conclusion, the human oxyntic epithelium is composed of continually renewing cells organized in distinct cell lineages.     

Structure-intestinal transport and structure-metabolism correlations of some potential cancerostatic pyrimidine nucleosides in isolated rat jejunum

Summary Both transport and biotransformation processes for a series of pyrimidine nucleobases, ribonucleosides, 2-deoxyribonucleosides, and acetyl and 5-substituted derivatives of the cancerostatic agent araC were studied in the isolated everted rat jejunum with a continuous perfusion technique. Metabolic alterations during penetration were assessed by HPLC. 5-Halogeno and 5-deoxy derivatives of cytosine nucleosides exhibited higher transport rates and higher stability towards the deamination reaction than did unsubstituted derivatives. Octanol-buffer partition coefficients were estimated for the study compounds, and fragmental constants for the sugar moieties of nucleosides were assessed. With the present study compounds there was no correlation between lipophilicity and transport rate, as previously reported, but there was a correlation between lipophilicity and metabolic alteration of araC derivatives (r=0.99, n=5).     

Teaching Case Ciliated Renal Tubular Cells in Crescentic Glomerulonephritis

Numerous cilia were found in the proximal convoluted tubules from a case of crescentic glomerulonephritis. The cilia exhibited the 9 + 2 pattern of organization characteristic of motile cilia. Microvilli were scanty or absent in heavily ciliated cells. The significance of renal cilia is discussed.     

Optimization of uretero-intestinal anastomosis in urinary diversion: an experimental study in dogs

Summary A new antireflux uretero-ileal reimplantation technique suitable for use with bladder substitutes is presented. This procedure entails creation of a serouslined extramural tunnel. Following detubularization of the bowel segment, the adjacent flaps are approximated by continuous 4/0 non-absorbable sutures 1.5 cm from the cut edges. The ureters are laid in the trough thus fashioned. Button-holes are created in the bowel flaps and a mucosa-to-mucosa uretero-ileal anastomosis is carried out. The mucosal edges of the flaps are then approximated by one layer of continuous 4/0 (PGA) suture resulting in closure of this artificial tunnel. The feasibility and functional outcome of this technique were experimentally investigated in 8 dogs. Follow-up was carried out up to 30 weeks. Assessment by intravenous urography and ascending studies showed that the procedure is an efficient method of providing an unobstructed unidirectional flow of urine.     

Aneurysms complicating pulmonary autograft procedure for aortic valve replacement

Aneurysm formation in the left ventricular outflow tract related to the proximal end of the pulmonary autograft after the Ross procedure was present in 2 patients. Both occurred late after operation and were associated with prolapse of a leaflet of the autograft and significant regurgitation. Both were repaired with no immediate complications. There was no evidence of infection at time of operation. The probable mechanisms underlying this complication and the possibilities of avoiding it are discussed.     

Correlation between fungi isolated from burn wounds and burn care units

A comparison was made prospectively between fungal isolates from patients and burn care units. Aspergillus niger was the most frequent isolate in both patients and burn care units whereas Ulocladium was the commonest isolate in the control group. Aspergillus terreus, Penicillium and Zygomycetes, which were recovered from burned patients, were also found more frequently in the burn care units than in the control group (other areas in the hospital). These findings indicate a potential risk of fungal infection which can be acquired from the immediate surroundings of patients in burn care units. Periodical burn ward decontamination is therefore recommended.     

Allied studies on the effect of Rosmarinus officinalis L. on experimental hepatotoxicity and mutagenesis

The hepatoprotective and antimutagenic effects of the rosemary essential oil and the ethanolic extract were investigated using carbon tetrachloride and cyclophosphamide as hepatotoxic and mutagenic compounds, respectively. Our results revealed that i.g. administration of the rosemary ethanolic extract (0.15 g/100 g BW) to rats for 3 weeks produced the most pronounced hepatoprotective effect compared to silymarin (reference compound) due to the amelioration of most of the studied serum and liver parameters and confirmed by histopathological examination of the liver tissue. Pretreatment of mice for 7 days with the rosemary essential oil (1.1 mg/g BW) followed by i.p. injection with cyclophosphamide reduced significantly the induced mitodepression in the bone marrow cells of the animals. The potential hepatoprotective and antimutagenic activities of the rosemary ethanolic extract and essential oil, respectively, are attributed to the presence of a relatively high percentage of phenolic compounds with high antioxidant activity (according to our chemical studies).