A simple method for monitoring changes in cell height using fluorescent microbeads and an Ussing-type chamber for the inverted microscope

In this study, we report two developments for studies of ion transport in cultured epithelial cells. First, a convenient method is presented for measuring apparent cell height using fluorescent microbeads as high-contrast landmarks of the apical and basal cell surfaces. The apparent cell height is then used as an indicator to monitor the time course of changes in cell volume in response to osmotic perturbations. Second, an Ussing-type chamber design for the inverted fluorescence microscope is presented, which allows determination of transepithelial electrical properties. Using these two methods, we obtained simultaneous measurements of cell height and transepithelial electrical parameters for cultured renal (A6) epithelium. Cell height was measured by alternately focusing the microscope between microbeads marking the apical and basal surfaces. The distance between these two surfaces was measured electrically from the voltage output of a potentiometer that was mechanically coupled to the fine-focusing knob of the microscope. Following decreases in the bathing solution osmolality, the cell height and transepithelial Na⁺ transport rate (measured as short-circuit current, I _SC) increased. The increase in cell height preceded changes in I _SC by several minutes, suggesting a lack of direct linkage between changes in cell volume and transepithelial Na⁺ transport. Both the fluorescent microbead cell height method and the Ussing-type chamber can be used in conjunction with patch-clamp techniques, intracellular microelectrode impalements, or fluorescent probes of intracellular composition. Therefore, this system may be advantageous for studies of epithelial cell volume and channel regulation.     

Cytologic grade independently predicts survival of patients with pancreatic adenocarcinoma

Our objectives were to devise a cytologic grading system and determine whether it would predict survival of patients with solid-type pancreatic adenocarcinoma. We evaluated 116 consecutive patients from July 2000 to November 2002; they were followed up until September 2003. We scored the following features on rapid Romanowsky-stained endoscopic ultrasound-guided fine-needle aspiration smears: cell group architecture, single cells, nuclear grade, mucus, bizarre cells, and necrosis. A cytologic grade (low vs high) was assigned. The Kaplan-Meier estimate of 6-month survival was 76% (SE, 7%) for patients with low-grade tumors vs 50% (SE, 6%) for patients with high-grade carcinoma. The median survival for patients with low-grade vs high-grade tumors was 1 year vs 6 months, respectively (chi2 = 4.45; P = .035). Cox proportional hazards regression showed tumor stage, cancer-specific treatment, and cytologic grade to be independent predictors of survival (P = .001). No other factors (age, mass location, placement of stent, presence of concomitant chronic pancreatitis, race, sex) predicted survival. We devised a grading system that independently predicted survival in patients with pancreatic adenocarcinoma.     

In Situ Thermal Denaturation of Proteins in Dunning AT-1 Prostate Cancer Cells: Implication for Hyperthermic Cell Injury

The in situ thermal protein denaturation and its correlation with direct hyperthermic cell injury in Dunning AT-1 prostate tumor cells were investigated in this study. The in situ thermal protein denaturation was studied using both Fourier transform infrared spectroscopy (FTIR) and differential scanning calorimetry (DSC). The FTIR spectra at different temperatures show changes in protein secondary structure (from alpha helix to extended beta sheet) during in situ thermal protein denaturation within AT-1 cells. Calorimetric studies using DSC show that endothermic heat release is associated with the in situ thermal protein denaturation. Furthermore, both the secondary structure changes detected by FTIR and the calorimetric changes detected by DSC were quantified and the kinetics of the overall in situ thermal protein denaturation was derived under different heating conditions. The onset temperature where the overall in situ thermal protein denaturation is first detectable was found to be scanning rate dependent (approximately 41 degrees C at 2 degrees C min(-1) and approximately 44 degrees C at 5 degrees C min(-1)). The kinetics of the overall in situ thermal protein denaturation was derived from both DSC and FTIR measurements and was fit using kinetic and statistical models. The kinetic data determined by FTIR and DSC under the same heating conditions match well with each other. The activation energy of the overall in situ thermal protein denaturation is found to be strongly dependent on the temperature range considered (the activation energy ranges from approximately 110 kJ mol(-1) between 44 and 90 degrees C to approximately 750 kJ mol(-1) between 44 and 50 degrees C). However, its dependence on heating rate is negligible. Several denaturation peaks, including a dominant one between approximately 62 and 65 degrees C, are identifiable from both the DSC and the FTIR results. To investigate directly the relationship between thermally induced cell injury and the in situ thermal protein denaturation, both acute (propidium iodide dye exclusion, assessed 3-h postthermal treatment) and chronic (clonogenics, assessed 7-day postthermal treatment) cell injury were quantified using AT-1 cells prepared under the same conditions as for the DSC protein studies. Comparisons of the results from the cell injury studies and the DSC protein denaturation studies show that the overall in situ thermal protein denaturation correlates well with both the acute and the chronic cell injury, which suggests that overall in situ thermal protein denaturation is an important mechanism of direct hyperthermic cell injury in AT-1 cells at the macromolecular level.     

The familial prevalence in second-degree relatives of patients with anxiety neurosis (panic disorder)

A family history study of second-degree relatives of 19 patients with anxiety neurosis (panic disorder) and 19 controls showed a morbidity risk of 9.5% among the former compared with 1.4% among the latter. These risks were approximately half those found among first-degree relatives. Female relatives were at higher risk for anxiety neurosis. The risk for other psychiatric illnesses did not differ between the relatives of anxiety neurosis and controls.     

Parental origin of de novo chromosome 9 deletions in del(9p) syndrome

Parental origin of de novo deletions in the short arm of chromosome 9 in patients with a clinical diagnosis of del(9p) syndrome was assessed in 13 patients using polymerase chain reaction (PCR) analysis of highly polymorphic dinucleotide repeat micro-satellite markers located in the putative deleted region. The deletion was found to be of paternal origin in 9 cases and of maternal origin in the remaining 4 cases, suggesting that the molecular event resulting in the deletion occurs in both male and female gametogenesis and that genomic imprinting does not appear to play a role in the patho-genesis of del(9p) syndrome. © 1995 Wiley-Liss, Inc.     

Drawbacks of GENEHUNTER for larger pedigrees: application to panic disorder

Large pedigrees can pose a problem for GENEHUNTER linkage analysis software. Differences in two-point and multipoint lodscores were observed when comparing GENEHUNTER to other linkage software. Careful consideration must be given when selecting linkage analysis programs. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:781-783, 2000.     

Bilateral microphthalmia with cyst,facial clefts,and limb anomalies: a new syndrome with features of Waardenburg syndrome,cerebro-oculo-nasal syndrome,and craniotelencephalic dysplasia

We report a patient with bilateral microphthalmia with cyst, limb anomalies, and multiple facial malformations. This patient has clinical features similar to Waardenburg ophthalmo-acromelic syndrome, cerebro-oculo-nasal syndrome, and craniotelencephalic dysplasia. Although all of these syndromes are characterized by microphthalmia, the presently reported patient does not have the complete pattern of any of these syndromes, It is possible that he has a previously undescribed syndrome, most closely related to the cerebro-oculo-nasal syndrome with malformations outside the craniofacial region. More case reports are needed to further delineate this possibly new syndrome.     

Culture-negative Bartonella endocarditis in a patient with renal failure: the value of molecular methods in diagnosis

Members of the genus Bartonella are increasingly recognised as a cause of culture-negative endocarditis, particularly in those patients with underlying risk factors (e.g., homelessness and alcoholism (B. quintana) or valvulopathy and cat ownership (B. henselae). The aortic and mitral-valves are most commonly involved. Here, a case is reported of culture-negative right-sided endocarditis, without any of the above risk factors, due to Bartonella sp. in a 69-year-old man who presented with acute renal failure. The diagnosis was made using a broad-range 16S rRNA polymerase chain reaction (PCR) technique and direct automated sequencing on a peripheral blood sample, which was subsequently confirmed serologically. A review of the literature on Bartonella endocarditis is also presented. Molecular laboratory methods using peripheral blood or blood cultures may be very useful in the diagnosis of causal agents in culture-negative endocarditis and add further support to the recently inclusion of molecular (PCR) diagnosis, as a major Duke's criterion, for the diagnosis of infective endocarditis.     

Single strand DNA breaks in mitogen stimulated T lymphocytes are religated by a mechanism independent of accessory cells.

The phenomenon of religation of single-strand DNA breaks (nicks) in mitogenically stimulated human T lymphocytes is an event occurring within 8 h of mitogen stimulation. Many later events in lymphocyte activation are known to be dependent on accessory cells, whereas earlier events are often accessory-cell independent. To establish whether nick religation is dependent or independent of accessory-cell function, lymphocytes were stimulated with PHA in the presence of inhibitors thought to act, in part at least, on accessory cells (methylprednisolone and cyclosporine A), or under conditions in which accessory-cell function is limited (low-density culture, adherent-cell depleted populations). In each case DNA synthesis was inhibited but the religation process was retained, indicating that it is independent of accessory-cell function. Inhibition of DNA synthesis in these cells was shown to be readily reversible on addition of conditioned medium containing accessory-cell products, but there was no further change in ligation.