Immunogenic Dendritic Cell Generation from Pluripotent Stem Cells by Ectopic Expression of Runx3.

Application of dendritic cells (DCs) to prime responses to tumor Ags provides a promising approach to immunotherapy. However, only a limited number of DCs can be manufactured from adult precursors. In contrast, pluripotent embryonic stem (ES) cells represent an inexhaustible source for DC production, although it remains a major challenge to steer directional differentiation because ES cell-derived cells are typically immature with impaired functional capacity. Consistent with this notion, we found that mouse ES cell-derived DCs (ES-DCs) represented less mature cells compared with bone marrow-derived DCs. This finding prompted us to compare the gene expression profile of the ES cell- and adult progenitor-derived, GM-CSF-instructed, nonconventional DC subsets. We quantified the mRNA level of 17 DC-specific transcription factors and observed that 3 transcriptional regulators (Irf4, Spi-B, and Runx3) showed lower expression in ES-DCs than in bone marrow-derived DCs. In light of this altered gene expression, we probed the effects of these transcription factors in developing mouse ES-DCs with an isogenic expression screen. Our analysis revealed that forced expression of Irf4 repressed ES-DC development, whereas, in contrast, Runx3 improved the ES-DC maturation capacity. Moreover, LPS-treated and Runx3-activated ES-DCs exhibited enhanced T cell activation and migratory potential. In summary, we found that ex vivo-generated ES-DCs had a compromised maturation ability and immunogenicity. However, ectopic expression of Runx3 enhances cytokine-driven ES-DC development and acts as an instructive tool for the generation of mature DCs with enhanced immunogenicity from pluripotent stem cells.

Value of carcinoembryonic antigen (CEA) and cholesterol assays of ascitic fluid in cases of inconclusive cytology.

AIM: To determine whether assays of carcinoembryonic antigen (CEA) and cholesterol in ascites add diagnostic value to cytology. METHODS: The additional diagnostic efficacy of the biochemical assays was studied in the ascitic fluid from 130 patients, of whom 57 had peritoneal carcinomatosis. All diagnoses were verified by subsequent necropsy and/or histology. RESULTS: CEA concentrations over 5 ng/ml indicated carcinomas, occasionally without peritoneal involvement of the tumour. However, increased values were significantly more common in cancer with peritoneal involvement (p < 0.01), giving a sensitivity of 51% and specificity of 97% for carcinomatosis. A cholesterol value exceeding 1.21 mmol/litre was found in 93% of cancers with peritoneal involvement, but it was not entirely specific (96%) for carcinomatosis. Simultaneous increases in CEA and cholesterol concentrations were specific for carcinomatosis and this combination increased the sensitivity for diagnosing carcinomatosis from 77% with cytology alone to 88%. The correct diagnosis could thus be made in five of 12 cases with inconclusive cytology. CONCLUSIONS: The measurements of both CEA and cholesterol concentrations in ascites give additional specific information about peritoneal carcinomatosis and can therefore be a useful adjunct to cytology-in particular, in inconclusive cases.

New human Dirofilarioses in Hungary.

About ten cases of filariosis have recently been recorded in the Hungarian medical literature, six of them caused by Dirofilaria repens. Dirofilaria repensis a mosquito-transmitted filaroid worm in the subcutaneous tissue of dogs and cats in the temperate areas of the Old World. It accidentally infects man, too, and can remain unidentified due to physicians, poor knowledge of the parasite. In the last two years six new Dirofilaria repensinfections have been found in various parts of the country: five localised dermally and one in the deep tissues. Two of the cases might have been acquired in Italy during summer travels. Four patients, however, have never been abroad, these cases must be considered autochtonous infections. The thickness of the multilayered cuticle of the worm, diameter of the body and the size, form and number of the longitudinal ridges on its surface are used in the histological diagnosis of the the parasite.

Central neurocytoma with malignant course. Neuronal and glial differentiation and craniospinal dissemination.

Central neurocytoma is a benign neuronal tumor of young adults in the lateral cerebral ventricles with characteristic X ray and light microscopic findings. In many respects typical central neurocytoma is reported below, with recurrence in the third month requiring reoperation. Death ensued in the fifth postoperative month. Subsequent histology proved progressive vascular proliferation and increasing, unusual glial differentiation of the neuronal tumor. At autopsy tumorous seeding blocked the liquor circulation. A thin tumorous layer covered the surface of all ventricles, the cerebellum and medulla oblongata. The GFAP positive cells out-numbered the synaptophysin positive ones. Increase of GFAP positivity and vascular proliferation of the central neurocytoma may be alarming signs suggesting a malignant course in addition to the other atypical features.

Human Dirofilaria repens infection in Hungary: a case in the spermatic cord and a review of the literature.

Orchiectomy was performed in a 37-year-old Hungarian man exhibiting a swelling in his right testicle. Histology revealed a nodule attached to the spermatic cord, consisting of a granulomatous tissue around sections of a nematode. The worm was identified as Dirofilaria repens, an uncommon parasite in Hungary. As the patient had been abroad only in Italy where cases of dirofilariosis in dogs and humans are relatively frequent, it is assumed that the infection might have been acquired in that country 5 years earlier. This is the fifth case, published so far in the world, of such a localization in a human. The human cases of dirofilariosis reported in Hungary are reviewed.

Histiocytic and t-cell rich b-cell lymphoma (TCRBCL) of the stomach.

Although stomach is a frequent site of extranodal lymphomas, histiocyte-rich TCRBCL has not yet been described there. Even histology of repeated gastrobiopsies of this uncommon, diffuse, large B-cell lymphoma may be inconclusive and partial gastrectomy cannot be avoided. It is only immunohistology (CD20, CD43, CD68) of the paraffin blocks from the resection specimen that can lead to the final diagnosis of intermediate grade malignant lymphoma.

Working experience with a new vacuum-accelerated microwave histoprocessor.

Methods of microwave and vacuum-accelerated fixation, dehydration, and paraffin embedding are described, using a new type of vacuum and temperature stabilizable microwave histoprocessor: MFX-800. The whole histoprocessing cycle lasts 3.5-5.5 h, depending on the thickness of the tissue blocks. Well-preserved structural detail, intense staining, and good antigen preservation were achieved with various tissues. This new histoprocessing facility is recommended for routine pathology laboratories in speeding up their processing procedures.

[Experiences with a new vacuum-accelerated microwave histoprocessor]. / Tapasztalataink egy új, vákuummal felgyorsított mikrohullámú hisztoprocesszorral.

Methods of microwave and vacuum accelerated fixation, dehydration and paraffin embedding are described. They can be carried out in a new type of vacuum and temperature stabilizable microwave histoprocessor: MFX-800. The whole histoprocessing lasts 3.5-5.5 hours depending on the thickness of the tissue blocks. Preserved structural detail, intensive staining and good antigen preservation were achieved with various tissues. This new histoprocessing facility can be highly recommended for pathology laboratories interested in speeding up the processing procedure, as well as in immunohistochemistry.

[Oncologic diagnosis of serous effusions in body cavities (cytologic study and evaluation of cholesterol and carcinoembryonic antigen levels)]. / A testüregi folyadékgyülemek daganatdiagnosztikája (A cytologia, a koleszterin és a carcinoembryonalis antigén vizsgálat együttes értékelése).

Conventional cytological evaluation of serous effusions often yields border line result: apart from positive (malignant) or negative (benign) diagnoses, a relatively large part of the findings are "suspicious for malignancy" (P3). In the present paper the authors have analysed to what extent contributes the determination of cholesterol and CEA levels of ascites and pleural effusion to the diagnostic accuracy of cytologically "suspicious" (P3) cases. In 155 histologically controlled cases, specificity, sensitivity and diagnostic efficiency were assessed on the basis of cytology as well as the determination of CEA and cholesterol levels. Statistical parameters were determined for each method separately and for their combined application. According to the findings, cholesterol and CEA levels over 1.16 mmol/l and 2.5 ng/ml, respectively, indicate malignancy. In 19 out of 29 cytologically "suspicious" cases (66%), which histologically proved to be positive, cholesterol and/or CEA levels were elevated. In all of the 12 non-neoplastic "suspicious" cases the two parameters were under the cutoff values. The application of an easy and inexpensive cholesterol test proves to be a sensitive technique for indicating carcinomatosis and it completes adequately the specific cytological evaluation. If clinical symptoms speak for tumor, but the cytology is negative, the evaluation of CEA level may prove to be useful as far as it can indicate cancer not yet accompanied by carcinomatosis.

An electron spin resonance (ESR) study of free radicals formed from hematoxylin on tissue sections after blueing (alkalization).

A free radical signal of 12 G width and g = 2.0045 can be observed in hematoxylin stained tissue blocks and sections. The amount of paramagnetic centres in stained specimens is significantly larger than in unstained ones. After alkalization simultaneously with the colour change the former free radical is detectable in hemalum precipitate and on stained paper strips. After solution of the stain in dioxane and alkalization, a well resolved hyperfine structure could be seen which could be assigned to three different radicals with the same g value as observed in the rigid matrix (tissue and paper). Quantitative evaluation of free radical concentration is also carried out for tissue sections.

A study of free radicals in paraffin embedded and deparaffinized human heart muscle tissue using electron spin resonance (ESR).

Free radicals (spectroscopic splitting factor; g factor = 2.003-2.005) were investigated in formol-fixed, paraffin embedded heart-muscle tissue sections using electron spin resonance (ESR) spectra. Changes in signal amplitude, g factor and line width were registered during deparaffinization, chloroform-methanol extraction, vapour treatment and bromination. An attempt was made to identify the source of the ESR signals by a correlation between the signal amplitude and number of fluorescent and/or Sudan-black-positive granules counted in the tissue sections. An increase in signal amplitude, g value and line narrowing were characteristic of the ascorbyl radical after deparafinization in air. Vapour treatment revelated that the broader signal has lower g factor, a characteristic that is tentatively assigned to oxidized lipids. The bromination resistant minor fraction of free radical centres with small g factor might be associated with the pigment content of the samples.

Large cell dysplasia of hepatocytes, is it a premalignant condition?

Incidence and character of large cell dysplasia (LCD) of liver were examined by histological, statistical and morphometric methods in an aged Middle European population. The necropsy material of 487 patients with micronodular cirrhosis and cirrhosis with primary liver tumour was studied. In addition, 50 primary tumours developed in non-cirrhotic livers, 34 metastatic tumours and 90 normal livers were examined. According to the findings, LCD often developed in aged patients. The high occurrence is presumably due to the high age. The frequency of LCD was significantly higher in active than in inactive cirrhosis. LCD could mostly be found around metastatic tumours, especially those with large, destructive foci. Sex differences occurred in a part of cases but an age dependence could not be demonstrated. Normal and dysplastic cells did not differ significantly in most morphometric parameters but there was a significant difference between tumorous and normal or dysplastic parameters. The data suggested that LCD is a secondary, reactive phenomenon occurring in various pathological processes.

Free radical signal of bile pigment in paraffin embedded liver tissue.

A linear correlation have been found between the amplitude of the free radical signal of the electron spin resonance (ESR) spectrum of paraffin embedded liver blocks and the number of bile casts in the histological sections made from these blocks. It has been suggested that the major part of the ESR free radical signal arises from bile pigment, but the contribution of "age pigment" cannot be excluded. On the basis of model experiments the majority of these radical centers was assigned to protein bound bilirubin. In the course of histological processing more than 80% of various free radical centers arising in air dried liver tissue is extracted.

The effect of E-N-L-trimethyllysine (TML) on the humoral and cellular immune response.

E-N-L-Trimethyllysine glutamate (TML) influences the humoral and cellular immune response of mice. Chronic pre- and post-treatment (100 mg/kg/day, 5 times) transitionally increased the anti-SRBC haemagglutinin titre of female CBA mice. After 400 r whole body irradiation, TML treatment accelerated the normalization of the haemagglutinin level. TML treatment prolonged the life-span of BDF1 hybrid mice that had first been immunized and then inoculated with L1210 cells. TML diminished the delayed type hypersensitivity reaction in vivo of irradiated and non-irradiated CBA female mice and dose-dependent decreased the spontaneous (SLMC) and antibody-dependent (ADCC) cytotoxicity of healthy human lymphocytes, in vitro. As a low molecular weight immunomodulant, TML may also be considered as a therapeutic tool.

The free radical signal of pigment gallstones.

Bile stones have been studied by Electron Spin Resonance (ESR) spectroscopy. Pigment containing stones show a 8.76 +/- 0.02 gauss wide free radical singlet at g approximately 2.003 region of the spectrum. Amplitude of this free radical signal linearly correlates with pigment content of the stone, which can be applied in the estimation of pigment content. The possible role of free radicals is suggested in the formation of pigment bile stones.

[Electron spin resonance study of melanin in paraffin-imbedded ocular melanoma specimens]. / Melanin elektron spin rezonanciás (ESR) vizsgálata paraffinba ágyazott okuláris melanomákban.

The Electron Spin Resonance (ESP) spectrum of ocular melanomas embedded in paraffin was studied. In the g approximately 2.003 range of the spectrum a mark of a free radical was detected. The correlation between the amplitude of the latter and the number of melanin granules in sections was studied. Considering the positive correlation revealed, authors believe, that ESR spectroscopy is a suitable tool for detecting melanin in paraffin embedded specimens. As an effect of oxidation or reduction melanin turns pale, however this phenomenon is not in tight connection with the decrease of the amplitude of the free-radical-mark. Very seldom different types of spectrum (peroxide, cupric ion) may appear, so it is reasonable to study large number of specimens.

ESR investigation of paraffin-embedded ocular melanomas.

Ocular melanomas embedded in paraffin wax for histological examination have been studied by electron spin resonance (ESR) spectroscopy. A free-radical signal was detected at the g = 2.003 section of the spectrum. The amplitude of this signal was correlated with the number of melanin granules in microscopical slides made from the tumour. This positive correlation can make ESR spectroscopy suitable for estimating the melanin content in the embedded melanoma blocks. Additional paramagnetic signals can also be detected. The clarification of their significance needs, however, further ESR measurements.

[E-N-trimethyl-lysine: a natural cellular component with mitogenic activity]. / E-N-trimetil-lisina: un componente cellulare naturale con attivita' mitogena.

The blastogenic capacity of a new mitogenic agent, E-N-trimethyl-lysine (TML) was examined on human peripheral blood lymphocytes. The results of combined TML-PHA treatment experiments suggested a difference in the trigger mechanism of these two mitogenic compounds.

epsilon-N-trimethyl-lysine: a natural cell component with mitogenic activity.

The blastogenic capacity of a new mitogenic agent, epsilon-N-trimethyl-lysine (TML) was examined on human peripheral blood lymphocytes. The changes in relative DNA and histone content after TML treatment testified that activated lymphocytes passed through the whole cell cycle. The comparison of the 3H-TdR incorporation rate into unseparated T-B lymphocyte and purified T lymphocyte cultures suggested a preferential effect on the thymidine incorporation of T lymphocytes. TML induced preferentially high responses in cases of blood cells taken from women. The possibility of DNA repair mechanism could be excluded. The results of combined TML-PHA treatment experiments suggested a difference in the trigger mechanism of these two mitogenic compounds.