A balancing act: orchestrating amino-truncated and full-length p73 variants as decisive factors in cancer progression.

p73 is the older sibling of p53 and mimics most of its tumor-suppressor functions. Through alternative promoter usage and splicing, the TP73 gene generates more than two dozen isoforms of which N-terminal truncated DNp73 variants have a decisive role in cancer pathogenesis as they outweigh the positive effects of full-length TAp73 and p53 in acting as a barrier to tumor development. Beyond the prevailing view that DNp73 predominantly counteract cell cycle arrest and apoptosis, latest progress indicates that these isoforms acquire novel functions in epithelial-to-mesenchymal transition, metastasis and therapy resistance. New insight into the mechanisms underlying this behavior reinforced the expectation that DNp73 variants contribute to aggressive cellular traits through both loss of wild-type tumor-suppressor activity and gain-of-function, suggesting an equally important role in cancer progression as mutant p53. In this review, we describe the novel properties of DNp73 in the invasion metastasis cascade and outline the comprehensive p73 regulatome with an emphasis on molecular processes putting TAp73 out of action in advanced tumors. These intriguing insights provoke a new understanding of the acquisition of aggressive traits by cancer cells and may help to set novel therapies for a broad range of metastatic tumors.

Transcriptome analysis in mouse tumors induced by Ret-MEN2/FMTC mutations reveals subtype-specific role in survival and interference with immune surveillance.

Activating mutations in the Ret proto-oncogene are responsible for occurrence of multiple endocrine neoplasia (MEN) type 2A and 2B, and familial medullary thyroid carcinoma (FMTC). A striking genotype-phenotype correlation between the mutated RET codon and clinical manifestation implies that tumorigenesis is conditioned by the type of mutation. We investigated gene expression profiles between and within distinct MEN2 subtypes through whole-genome microarray analysis in tumors induced by NIH-3T3 cells transformed with defined RET-MEN2A (C609Y, C634R), MEN2B, (A883F, M918T), and FMTC (Y791F) mutations. Expression profiling identified a statistically significant modification of 1494 genes, 628 down- and 866 upregulated in MEN2B compared with MEN2A/FMTC tumors. By contrast, no obvious alterations were observed among individual MEN2B and MEN2A type mutations, or between MEN2A and FMTC. Functional clustering of differential genes revealed RET-MEN2B specific upregulation of genes associated with novel growth and survival pathways. Intriguingly, RET-MEN2A/FMTC-specific tumors were characterized by a considerable number of genes involved in the host antitumor immune response via stimulation of natural killer/T-cell proliferation, migration, and cytotoxicity, which were completely absent in RET-MEN2B related cancers. QPCR on tumors versus cultured NIH-RET cell lines demonstrated that they are largely attributed to the host innate immune system, whereas expression of CX3CL1 involved in leukocyte recruitment is exclusively RET-MEN2A/FMTC tumor cell dependent. In correlation, massive inflammatory infiltrates were apparent only in tumors carrying MEN type 2A/FMTC mutations, suggesting that RET-MEN2B receptors specifically counteract immune infiltration by preventing chemokine expression, which may contribute to the different clinical outcome of both subtypes.

[Quality control of thoracic X-rays in children in diagnostic centers with and without pediatric-radiologic competence]. / Qualitätskontrolle von Röntgenthoraxaufnahmen bei Kindern in diagnostischen Zentren mit und ohne kinderradiologische Kompetenz.

PURPOSE: Radiation protection in pediatric radiology is very important because of the particular sensitivity of radiosensitive organs in younger patients. Optimized image quality supports radiation protection and should be targeted. In our study we examined the quality of pediatric chest X-rays at diagnostic centers (university hospitals and other large clinics). We then evaluated differences in image quality in departments without pediatric competence (R) and departments with pediatric competence (PR). MATERIALS AND METHODS: Our study was based on 313 conventional chest X-rays from 207 patients (192 p. a./a. p. and 121 lateral, 43 from R, 258 from PR and 12 neither from R nor KR) and 38 digital chest X-rays from 26 patients (25 p. a./a. p. and 13 lateral, 1 from R and 37 from PR). All patients (age 0 - 18 years) are from Nephroblastoma-Study SIOP-93/01-GPOH. We examined all initial chest X-rays, which were sent to us for evaluation upon request between 4/3/2002 and 6/14/2002. The examined parameters were: exposure, centering of the X-rays/patient positioning, collimation and sharpness. The X-rays were evaluated on a scale from 1 (best result) to 5 (worst result), resulting in an overall score of A = optimum, B = minor problems, C = major problems, or D = unusable. The optical density, the center of the image and the relative field size were also measured. Statistical tests (Mann-Whitney-U and log regression) were carried out on the conventional images. The study was performed retrospectively. The exposure, sharpness and optical density of the digital X-rays were not analyzed. RESULTS: In the case of all conventional X-rays, the quality of the centering of the X-rays/patient positioning and collimation was moderate (average scale value: 2.4 and 2.8), and the quality of the exposure and sharpness was good and very good (average scale value: 1.9 and 1.5). The quality of the chest X-rays in departments with additional pediatric radiological expertise was better mainly in the case of younger patients (younger than 5 years) than departments without additional pediatric radiological expertise (average scale value in age group 0 - 1 month: PR = 1.7; average scale value in age group 2 months - 2 years: R = 2.4 and PR = 1.8; average scale value in age group 3 - 5 years: R = 2.5 and PR = 1.8). CONCLUSION: Despite the good overall image quality, the quality of the centering of the X-rays/patient positioning and collimation was insufficient in both examiner groups (R and PR). For this reason, some radiation protection requirements could not be fulfilled. X-rays from PR were higher quality than X-rays from R in this special study group. Day-to-day quality checks are necessary for pediatric chest X-rays in order to achieve a high quality standard.

[Radiologic differentiation of rhabdoid tumor from Wilms' tumor and mesoblastic nephroma]. / Bildgebende Differenzierung des Rhabdoidtumors vom Nephroblastom und mesoblastischen Nephrom.

Differentiation between rhabdoid tumor (RT) and mesoblastic nephroma (MN) and Wilms' tumor (WT) by imaging studies in babies and young children before histological confirmation is useful to start optimal treatment early. Typical radiologic criteria (crescent-shaped subcapsular liquid areas, tumor lobules, blurred tumor borders, metastasis in the lung, and regional lymph nodes) are described. The results of 26 MRI, 30 CT, and 22 ultrasound examinations of 49 patients (22 RT, 19 WT, and 8 MN, age 2-57 months) were analyzed. The above-mentioned radiologic criteria were classified with score values. The score value distribution was analyzed between the tumor entities and by two investigators.RT had significantly higher score values than the MN and WT. The difference between the two investigators was not significant. As a group RT differentiates from the group of WT and MN, but this is not possible in single cases with the radiologic criteria employed. Only if more signs are observed together in one case can a RT be presumed, which may indicate an early biopsy before chemotherapy.

[Rhabdoid tumors of the kidney in childhood]. / Rhabdoidtumoren der Niere im Kindesalter -- Eine retrospektive radiomorphologische Analyse von 22 im Rahmen der Nephroblastomstudie SIOP 93/01-GPOH registrierten Fällen.

PURPOSE: The primary diagnosis of renal masses in children is made by imaging studies. This retrospective analysis describes the imaging features of rhabdoid tumors (RT) with US, CT and MRI, to point out characteristics and to evaluate the possibility of differentiation between RT and Wilms tumor. MATERIALS AND METHODS: We reviewed 10 MRI (6 STIR, 9 T1 w, 8 T2 w, 10 T1 post KM), 15 CT (9 Nativ-CT, 14 KM-CT) and 14 US images of 22 patients (age 2 - 57 months) with histopathologically confirmed RT. The following characteristics were evaluated: subcapsular fluid collection, multiple tumor lobules, presence of calcification, primary tumor size, visibility of tumor margin, tumor necrosis and metastases. RESULTS: The mean total tumor volume was 238 ml. 19 RT were located in the perihilar/medullary region with invasion of the renal hilum, and 5/22 tumors showed multiple tumor lobules. Subcapsular fluid collection was found in 6/22 cases. Calcifications were present in 6/19. Eleven tumors were well defined from the renal parenchyma, 9 poorly defined, 2 could not be assessed. In 19/22 cases tumor necrosis was found. Distant metastases were seen in 8 cases in the lung, in 3 cases in the CNS. Metastases of regional lymph nodes were seen in 9 cases. CONCLUSION: The evaluated characteristics frequently found in RT are not indicative of these tumors. RT cannot clearly be differentiated from Wilms tumor by imaging studies. Because of frequent involvement of the CNS and lung, a MRI of the CNS and CT of the lung is indicated after histopathologic diagnosis of RT is made.

Effects of feeding high dosages of vitamin E to laying hens on thyroid hormone concentrations of hatching chicks.

Long-term experimental feeding of 20,000 ppm alpha-tocopheryl acetate to laying hens caused a significant (P < 0.05) decrease in hatching rates as compared to the control group, which was fed a diet containing 19 ppm alpha-tocopherol. When the thyroid hormones in the developing chicks were checked on incubation days 16, 19, 21, and 22, the following results were ascertained: During the latter part of incubation, increases in plasma concentrations of thyroxine and triiodothyronine were observed. No significant differences in hormone concentrations (P > 0.05) between the control and the treatment group were observed during incubation days 16, 19, and 22. However, on the day of hatching (day 21 of incubation) significantly lower (P < 0.05) triiodothyronine concentrations in chick embryos of piped eggs were found in the treatment group. Moreover, thyroxine concentrations in non-piped eggs and in hatched chicks were found to be significantly higher as compared to the control group. Given these results, one concludes that extremely high dosages of vitamin E may affect thyroid hormone concentrations of hatching chicks, and therefore, the chicks might be inhibited in pipping the egg shell. Hypothetically, the hepatic enzyme 5'-monodeiodinase is involved in the mechanism of inhibition.

[Quality of ambulatory thoracic radiography in the child--a pilot study]. / Qualität der ambulanten Röntgenthoraxuntersuchung des Kindes--eine Pilotstudie.

A quality control of outpatient paediatric chest X-rays was conducted in a sample of patients of one paediatric practice. During a period of eight months the technical image quality was analysed considering both diagnostic aspects and radiation protection. The quality of the 139 examined chest X-rays was inadequate concerning the collimation and focussing of the X-rays and the positioning of the patients. Exposure was estimated as average, sharpness was rated as good. In total 14% of the X-rays were not suitable for medical diagnosis. Image quality of the X-rays of infants (children younger than 6 years) was significantly lower compared to the total sample. Radiation protection standards were not fulfilled. As a conclusion from our results, improvements in outpatient paediatric radiography are urgently necessary. Quality control committees should pay particular attention in radiographs of infants.

The glycosaminoglycans in cultures of stimulated rat peritoneal macrophages. 2. Gel chromatographic studies and the behaviour of heparan sulfate.

The molecular weight distribution of pMP-derived glycosaminoglycans (GAG), i.e. non-sulfated GAG, chondroitin sulfate (CS), and heparin sulfate (HS)-like material was determined. The peritoneal macrophages (pMP) were harvested from rats normal or stimulated by i.p. injection of thioglycolate, carrageenan or BCG, and maintained in culture. The GAG of cell layer and medium were isolated separately after labeling with 35S-sulfate and 3H-acetate. Treatment with nitrous acid served to remove HS-like material. Labeling with 3H-acetate served to detect synthesis of the high m. w. hyaluronic acid (HA). Gel chromatic separation was done using Sephadex G-200 columns. The maximal size of 35S-labeled GAG, especially HS (36 kDa), was reduced in cultural medium and cell layer after stimulation in vivo. Reduction was most pronounced after application of carrageenan followed by thioglycolate and BCG/LPS stimulation. The extracellular GAG of BCG-stimulated pMP were smallest, probably due to degradation. Heparan sulfate-like material made up a larger proportion in monolayer and medium, comprising the total m.w. range up to 36 kDa. The GAG sensitive to nitrous acid were maximal in cultures of carrageenan-stimulated pMP and minimal in those of thioglycolate-stimulated pMP. This type of HS was sensitive to hyaluronidase, too. Any synthesis of high molecular hyaluronic acid was not found in normal or stimulated rat pMP. Therefore MP-associated HA must be adsorbed from other sources or synthesized by early forms of macrophages.

The glycosaminoglycans in cultures of stimulated rat peritoneal macrophages. 1. Pattern and biosynthesis of glycosaminoglycans.

Macrophages produce and secrete proteoglycans. They are involved in inflammation and may contribute to the glycosaminoglycans (GAG) and proteoglycans (PG) characteristic of the inflamed area. This possible contribution was studied with rat peritoneal macrophages (pMP) in vitro. The total amount, composition, and neosynthesis of the GAG (as the main constituents of PG) were determined in cultured pMP of normal rats or rats pretreated with casein, BCG, thioglycolate or carrageenan. Partly the stimulated pMP were further activated with LPS or PMA. The rat pMP contained non-sulfated GAG, chondroitin sulfate and heparan sulfate but no dermatan sulfate. The GAG amount per cell increased with duration of the culture. In most cases the final level of GAG roughly corresponded to that of 12 micrograms hexuronic acid per 10(6) cells. It was lower after BCG stimulation (25%) and higher in pMP stimulated by both casein and LPS (200%). The average percentage of the GAG secreted (40%) was enhanced with pMP stimulated by carrageenan (87%) or casein plus LPS (69%). The pattern of the GAG in cultural media, cell coat and cells differed markedly. Thus the cell coat contained a higher amount of heparan sulfate. Differences due to stimulation were mainly seen in the reduced sulfation of the CS-proteoglycan secreted (casein-, BCG-pMP). According to studies on the incorporation of 35S-sulfate and 3H-acetate the GAG of the cell coat are less labeled and more conservative. The bulk of newly synthesized GAG/PG is secreted. This secretion appears an important property of macrophages the cause of which is speculative.

The glycosaminoglycan metabolism of chondrocyte monolayer cultures under normal and pathological conditions. A methodic study.

Chondrocyte cultures may serve as a model in investigating changes of the cartilage metabolism. Adherent chondrocytes in vitro maintain polygonal morphology at high cell density in the primary and secondary culture. Collagen type II is only clearly detected in multilayered or nodular areas. The differentiation of the chondrocytes is also indicated by a low HA concentration of the cultural medium. It depends on high cell density, a low number of subcultures and their duration. However, the medium GAG of chondrocyte cultures does not exactly mirror the state of cell differentiation but can partly be used to check it. Subcultures of chondrocytes on small cover slides (minicultures) are used to determine proteoglycan synthesis and degradation for 48 h each. Both synthesis and degradation of cell-associated GAG or proteoglycans, resp., follow similar complex kinetics. The half lives of sulfated GAG or proteoglycans are initially 10 h (T-1 for O-6 h of chase), later 39 h or 95 h (T-2 for 6-48 h of chase). Conditioned medium of casein-elicited rat peritoneal macrophages reduce the sulfate incorporation into chondrocyte proteoglycans and their degradation rates increase. In the additional presence of E. coli endotoxin (0.5 microgram/ml) the synthesis of proteoglycans is only little affected; the degradation rate is stronger increased. To peritoneal macrophages of rats manifold pretreated with BCG and perhaps desensitized, LPS is added in vitro. Conditioned medium of these MP does not affect the chondrocyte proteoglycan synthesis but enhances the degradation rates in a concentration-dependent manner. Thus it can be demonstrated that chondrocyte monolayer miniscale cultures may serve to elucidate changes in the proteoglycan synthesis and different degradative steps.

Bacillus Calmette-Guérin (BCG) influences cell proliferation and glycosaminoglycans of chondrocyte cultures.

There are only few reports on the correlation between bacterial products and the GAG pattern of cartilage. Mycobacteria bovis (BCG) were applied to chondrocyte monolayer cultures for one week. The following parameters did change: cell proliferation increased, glycosaminoglycan synthesis and secretion decreased, hyaluronic acid in secreted and cell-associated glycosaminoglycans increased, a correlation between the degree of these changes and the degree of cell differentiation seems to exist. The contact of bacteria like BCG to chondrocytes may change the cellular metabolism. On the tissue level this may injure articular cartilage and thus support the concept of predamaged cartilage that is readily susceptible to further degradation.

Effect of the microbial constituents, LPS and BCG, on the glycosaminoglycans of chondrocyte cultures.

Earlier we reported that articular chondrocytes in monolayer culture produce pericellular proteoglycans both with short and long half lives, T-1 and T-2 (Kittlick et al. 1991 b). Now monolayer cultures have been investigated to assess the influence on the metabolism of pericellular proteoglycans or glycosaminoglycans by lipopolysaccharide of E. coli and S. typhimurium as well as by Bacillus Calmette-Guérin.

The glycosaminoglycans of the macrophage-like cell line PeMa in suspension and monolayer culture.

The macrophage-like cell line PeMa was cultured both in suspension and in monolayer for biochemical studies. It is concluded that adherent PeMa when compared with suspended cells --contain three times more GAG in medium, cell coat, and cells: --contain higher amounts of HS and CS in the cell coat; --release a minor proportion of sulfated GAG (HS) into the medium; --contain in their medium longer chains of HS and CS/DS; --contain in cell coat and cells higher amounts of a family of HS chains with high to low molecular weights. In suspension and monolayer cultures, the cell coat GAG were scarcely labelled; they are likely to be produced at another time and to turn over slowly. It is suggested that this study will be helpful to understand the behaviour of macrophages and other cells.

Modification of fibroblastic glycosaminoglycan pattern by soluble factors obtained from sensitized lymphocytes.

Lymph node cells were obtained from BCG-sensitized guinea pigs and cultured without (C) and with (Ag) PPD challenge. The dialyzed lymphocyte (LC) supernatants were added to the medium of monolayer cultures of embryonic rat fibroblasts. They were assayed at different cell densities and in the presence and absence of serum. The following results were obtained: Glucose consumption was increased but the cell counts were reduced with both the types of LC supernatants, suggesting a cytotoxic effect. Small increases of cell counts were observed with Ag supernatant in high density fibroblasts in absence of serum. Glycosaminoglycan levels per cell in medium and monolayer were similarly enhanced by both the types of supernatants in presence of serum. As an exception, Ag supernatant reduced cell glycosaminoglycans (GAG) at low cell density. In the absence of serum at low cell density, the increase of medium and cell GAG was markedly higher with Ag supernatant. The GAG pattern of the fibroblast media was scarcely influenced by LC supernatants. In the cell monolayer, however, interesting changes have been observed. Increases of HS and DS were produced by C-supernatant in low density fibroblasts as well as by C- and Ag-supernatants in high density fibroblasts with serum. HS was also increased by C- and Ag-supernatants in serum-free low density fibroblasts. Increases of HA and CS were produced by Ag supernatant in low density fibroblasts with serum. The possible significance of these changes is discussed with regard to chronic inflammation.