Visualizing metabolic changes in breast-cancer tissue using 1H-NMR spectroscopy and self-organizing maps

In-vitro NMR spectroscopic examinations of tissue extracts can be combined with appropriate pattern-recognition and visualization techniques in order to monitor characteristic metabolic differences between tissue classes. In the present study, such techniques are applied to a set of 88 breast-tissue samples with the intention of identifying typical differences between various tissue classes. The set contains 49 breast-tumor samples of various tumor grades and 39 samples of healthy tissue. The metabolite compositions of the tissue extracts were investigated using a dual extraction technique and high-resolution (1)H-NMR spectroscopy. The spectra of the hydrophilic and the lipophilic compounds were assigned to three groups according to different malignancy grades of the respective tissue samples. The group characteristics were analyzed using the k-nearest-neighbor method and self-organizing-map visualizations. The results show an increase of UDP-hexose, phosphocholine and phosphoethanolamine concentrations according to the tumor grade. Higher concentrations of taurine were detected in the malignant samples. Myo-inositol and glucose content were elevated in control samples compared with malignant tissue. Both compounds also characterized different subgroups in the pool of unaffected tissue samples depending upon fat content or fibrosis. Several lipid metabolites showed a characteristic elevation with high malignancy.     

DNA end-independent activation of DNA-PK mediated via association with the DNA-binding protein C1D

DNA-dependent protein kinase (DNA-PK), which is involved in DNA double-strand break repair and V(D)J recombination, is comprised of a DNA-targeting component termed Ku and an ~465-kD catalytic subunit, DNA-PK_cs. Although DNA-PK phosphorylates proteins in the presence of DSBs or other discontinuities in the DNA double helix in vitro, the possibility exists that it is also activated in other circumstances via its association with additional proteins. Here, through use of the yeast two-hybrid screen, we discover that the recently identified high affinity DNA binding protein C1D interacts with the putative leucine zipper region of DNA-PK_cs. Furthermore, we show that C1D can interact with DNA-PK in mammalian cells and that C1D is a very effective DNA-PK substrate in vitro. Finally, we establish that C1D directs the activation of DNA-PK in a manner that does not require DNA termini. Therefore, these studies provide a function for C1D and suggest novel mechanisms for DNA-PK activation in vivo.     

Mineralization behaviour of collagen type I immobilized on different substrates

Collagen type I as a robust fibre protein and main component of the extracellular matrix of most tissues is increasingly utilized for surface engineering of biomaterials using different immobilization methods. In the present work we studied the mineralization behaviour of fibrillar collagen type I in simulated body fluid as a measure for conformational changes caused by adsorptive immobilization or immobilization by partial incorporation into the anodic oxide layer on c.p.-titanium using microscopic and vibration spectroscopic methods. Adsorptive immobilization on highly oriented pyrolytic graphite (HOPG) and c.p.-titanium without collagen were used as references. In the initial phase (1-24 h) the kinetics of formation and the morphology of calcium phosphate phases (CPP) are strongly influenced both by the substrate and the immobilization method. Compared to HOPG both types of immobilization on titanium increasingly inhibit the formation of CPP. For longer times (30 d) these initial differences disappear-mineralization product on titanium, irrespective of the presence of collagen, is a mixture of amorphous calcium phosphate and octacalcium phosphate. Contrary to this the mineralization of HOPG substrates results in hydroxy apatite. This is discussed with respect to the conditions during the immobilization as well as the resulting interactions between substrate and immobilized collagen. It is shown that the mineralization process exhibits a high sensitivity with respect to conformational changes caused by these interactions. Possible cell biological relevance of these conformational changes is discussed.     

Molecular and cytogenetic characterization of a non-mosaic isodicentric Y chromosome in a patient with Klinefelter syndrome

We report on an adult male with Klinefelter phenotype and an isodicentric Y chromosome (47,XX,+idic(Y)(q12)), a combination which has to the best of our knowledge not been reported before. The patient was hospitalized in forensic psychiatry because of repeated delinquency, aggressive, aberrant and inappropriate behavior, and borderline intelligence. Molecular cytogenetic studies (FISH) showed that the SRY gene was present on both ends of the idicY, while there was only one signal for the Yq subtelomere probe. Molecular investigations by multiplex PCR, using STS markers covering the short and long arm of the Y chromosome did not indicate a deletion of Y chromosomal material. Molecular investigations of STR markers located on Xp22.3 and Xq28 indicated paternal origin of the additional X chromosome and an error in paternal meiosis I. Results of FISH analysis and molecular investigations are compatible with a phenotype as described for individuals with a 48,XXYY karyotype and support the findings that isodicentric Y chromosomes are frequently accompanied by other sex chromosomal abnormalities.     

In vivo labeling with1251-CRH of human ACTH-producing pituitary adenomas heterotransplanted to nude mice

Tissue from 23 pituitary adenomas causing Cushing’s disease was implanted subcutaneously into 159 NuNu/NMRi mice, resected after 21 or 35 days, and evaluated histologically and immunohistochemically. After 21 days, 74.3% of the grafts survived, 59% having less than 30% necrotic adenoma cells. After 35 days, 45% of the adenoma fragments survived, 37% having less than 30% necrotic adenoma cells. The preservation of the grafts was essentially dependent on the grade of vascularization accomplished by migration of the host’s capillaries. As assessed by adrenal weight and histologically, biological activity of the transplants could not be detected. Histologically, the grafts maintained the features of their primary tumors, and adrenocorticotropic hormone (ACTH) could be visualized immunohistologically.Seventeen mice with subsequently proved preserved adenoma tissue received an intravenous injection of 12.5 μCi¹²⁵l-corticotropin-releasing hormone (CRH) and light microscopy-autoradiography was performed. Specific labeling, as verified by positive and negative controls, was exhibited by 1 1 of 15 transplants originating from 3 highly differentiated ACTH cell adenomas. Four did not label clearly positive. Two grafts of an undifferentiated mucoid cell pituitary adenoma did not show any labeling.The nude mouse model is a useful tool for the study of ACTH-producing pituitary adenomas in vivo. Highly differentiated ACTH cell adenomas can be labeled with radioactive CRH in vivo.     

Amygdala-prefrontal coupling depends on a genetic variation of the serotonin transporter

Major depression is conditionally linked to a polymorphism of the human serotonin transporter gene (SLC6A4). During the presentation of aversive, but not pleasant, pictures, healthy carriers of the SLC6A4 short (s) allele showed stronger activation of the amygdala on functional magnetic resonance imaging. s carriers also showed greater coupling between the amygdala and the ventromedial prefrontal cortex, which may contribute to the abnormally high activity in the amygdala and medial prefrontal cortex seen in major depression.     

Red cell hemoglobin,hydrogen ion and electrolyte concentrations during exercise in trained and untrained subjects

Summary Red cell concentrations of hemoglobin (MCHC), H⁺, Na⁺, K⁺, Mg⁺⁺, Cl^– were measured in femoral venous blood of six untrained (UT), six endurance trained (TR) and three semitrained (ST) subjects during graded increasing work (4, 8, 12, 18 and 24 mkp/s, 10–15 min on each step) on a bicycle ergometer. Before exercise no significant differences were detected for the measured variables when comparing UT and TR. During exercise MCHC, [Na⁺], [K⁺] and [Mg⁺⁺] remained constant indicating lack of water shift into the erythrocytes in spite of a marked acidosis (lowest pH_Blood value 7.225). This lack resulted from an elevated extracellular osmolality. [H⁺]_Ery and [Cl^–]_Ery maximally increased by 2.0×10^–8 eq/kg H₂O and 10 meq/l, respectively. The change was markedly greater in UT than in TR at equal load. However, if [H⁺]_Ery and [Cl^–]_Ery were related to pH of whole blood, differences between groups almost disappeared and the ions were distributed as predictable from in vitro experiments (Fitzsimmons and Sendroy, 1961). Behaviour of H⁺ and Cl^– may be of importance for oxygen dissociation under in vivo conditions.Supported by Bundesinstitut für Sportwissenschaften, Köln     

Predicting the development of infant emotionality from maternal characteristics

Few studies have examined the associations between environmental conditions and developing infant emotionality or the differential susceptibility to those conditions. The present longitudinal study aims to make a contribution to close that gap. We analyzed whether positive emotionality, negative emotionality/irritability, and withdrawal/fear at the end of the first year of life are predictable from preceding caregiver's depression/anxiety, social support, and sensitivity in the interaction with the infant while controlling for antecedent states of emotionality. Furthermore, the question of whether associations between maternal characteristics and subsequent fear are stronger in the subgroup of infants high in irritability as opposed to those who are low in irritability was investigated. Subjects were 101 healthy firstborn infants and their primary caregivers. Assessments were conducted at infant ages of 4, 8, and 12 months. Depression, anxiety, and the social support of the caregiver were assessed by questionnaire. Sensitivity in the caregiver-infant interaction was assessed by behavior observations within the scope of home visits. Temperament characteristics were observed in standardized laboratory episodes. Whereas negative emotionality and withdrawal/fear were significantly predictable from the maternal characteristics, no predictability could be shown for developing positive emotionality. There were indications of a stronger association between the maternal characteristics and developing withdrawal/fear in irritable infants.