Biomedical subjects
Publications and source records attributed to A Klein.
The technique of physical match, which is widely used with materials such as plastic and glass, is applied to fragmented bullets for determination of common origin. The more conventional method, of tool marks comparison, is later used to connect the bullet with a firearm.
BACKGROUND: Cytokeratins are constituents of the intermediate filaments of epithelial cells that are expressed in various combinations depending on the epithelial type and the degree of differentiation. The recently identified cytokeratin 20 (CK-20) was found to be expressed in colonic, gastric, and pancreatic carcinoma tumor tissues. A low rate of incidence of expression of CK-20 was found in tumor tissue from lung carcinoma but no expression was found in blood even with the sensitive reverse transcriptase-polymerase chain reaction (RT-PCR) method. The objective of the current study was to examine whether CK-20 expression in the blood can be used as a biomarker for the detection of dissemination in patients with carcinoma of the colon, stomach, and pancreas. METHODS: In the current study, RT-PCR was used to determine the expression of CK-20 in the blood cells from patients with metastatic colon carcinoma (n = 22), metastatic pancreatic carcinoma (n = 28), metastatic gastric carcinoma (n = 18), metastatic lung carcinoma (n = 13), no metastatic colon carcinoma (n = 13) and no known malignant diseases (n = 22). RNA was extracted from cell pellets and analyzed by RT-PCR using primers for CK-20. RESULTS: In the group of 22 patients with metastatic colon carcinoma, 14 were found to be CK-20 positive (sensitivity of 63.6% and specificity of 92.3%), 22 of the 28 pancreatic carcinoma patients showed positive CK-20 expression, and 12 of 18 patients with gastric carcinoma showed positive CK-20 expression. All patients with metastatic lung carcinoma except 1 were negative (12 of 13 patients), and 12 of 13 patients with colonic carcinoma with no known metastases also were negative. Negative CK-20 results were obtained in all 22 patients with no known malignant diseases. CONCLUSIONS: The results of the current study indicate that because of its high sensitivity, RT-PCR of CK-20 is a potential biomarker for detecting metastases in blood samples from patients with carcinoma of the colon, stomach, and pancreas.
Glycoproteinoses belong to the lysosomal storage disorders group. The common feature of these diseases is the deficiency of a lysosomal protein that is part of glycan catabolism. Most of the lysosomal enzymes involved in the hydrolysis of glycoprotein carbohydrate chains are exo-glycosidases, which stepwise remove terminal monosaccharides. Thus, the deficiency of a single enzyme causes the blockage of the entire pathway and induces a storage of incompletely degraded substances inside the lysosome. Different mutations may be observed in a single disease and in all cases account for the nonexpression of lysosomal glycosidase activity. Different clinical phenotypes generally characterize a specific disorder, which rather must be described as a continuum in severity, suggesting that other biochemical or environmental factors influence the course of the disease. This review provides details on clinical features, genotype-phenotype correlations, enzymology and biochemical storage of four human glycoprotein lysosomal storage disorders, respectively alpha- and beta-mannosidosis, fucosidosis and alpha-N-acetylgalactosaminidase deficiency. Moreover, several animal disorders of glycoprotein metabolism have been found and constitute valuable models for the understanding of their human counterparts.
Macronuclear chromosomes of hypotrichous ciliates are gene-sized molecules carrying the coding sequence flanked by short non-translated regions and bounded by telomeres. We have constructed artificial chromosomes for investigation of transcription in the macronucleus of Euplotes crassus. The neo gene was put under the control of the 5"-non-translated region of the TBP gene of E.crassus. These molecules were introduced into the cell with the help of liposomes. The cells were transformed and survived high concentrations of geneticin. The artificial chromosomes were kept in the macro-nucleus for at least 50 days at a copy number of about 200 per macronucleus. Expression of the gene was shown by reverse transcription of the neo messenger. The transcription start was mapped and found to coincide with that found on the natural macronuclear chromosome encoding TBP in E.crassus.
Cognitive impairment is common in patients with advanced liver disease. It has been suggested that patients with alcoholic liver disease (ALD) have more impaired cognition than nonalcoholics. The objective of this study was to characterize any differences in cognitive functions between alcoholic cirrhotic patients and non-alcoholic cirrhotic patients of similar age, education, and severity of liver disease. We assessed cognitive functions in 117 patients with alcoholic cirrhosis and 163 patients with nonalcoholic cirrhosis using a brief battery of neuropsychological tests. In addition, all patients had standard psychiatric examinations to assess the effect of the disease severity, alcoholism, anxiety, and depression on the test scores. The study showed a higher proportion of patients with cognitive impairment in the alcoholic group. Alcoholics performed poorly in tests of memory and motor speed compared with nonalcoholics, despite similar premorbid IQ and education. Because patients with alcoholic cirrhosis had more severe liver disease (Child-Pugh score 8.5 +/- 2.2 vs. 7.6 +/- 2.2, P =.03) than nonalcoholics, the results were reanalyzed after adjusting for the linear effects of Child-Pugh score on cognitive test scores. We also used two-way analysis of variance to examine the interaction between Child class and alcoholism. Finally, the test scores were compared within each Child class. These analyses revealed no primary or interaction effect of alcoholism and confirmed that the differences in the test scores observed in alcoholics reflect the greater severity of their liver disease. The severity of cognitive impairment is similar in both alcoholic and non-alcoholic cirrhotic patients when adjusted for the severity of liver disease.
Tacrolimus is an effective immunosuppressant in the rescue of liver allograft patients in whom conventional immunosuppression failed. Efficacy and safety were examined in a multicenter trial of liver transplant recipients converted to tacrolimus because of rejection despite cyclosporine (CyA) therapy or intolerance to CyA. Six hundred seventy-seven patients were enrolled onto the study; 475 patients for rejection, 197 patients for intolerance, and 5 patients treated compassionately. The mean daily dose of tacrolimus was less in the intolerance (Int) patients throughout the study: 0.22 versus 0.17 mg/kg at 1 week and 0.14 versus 0.11 mg/kg at 24 months in rejection (Rej) and Int patients, respectively. Mean blood levels paralleled dosing in both groups, but were greater in the Rej patients (10.7 v 8.3 ng/mL at 18 months). Kaplan-Meier estimates of patient and graft survival were similar in the two groups. Patient survival rates were 80.1% and 81.5%, and graft survival rates were 72.7% and 73.9% at 24 months in the Rej and Int patients, respectively. Most adverse events occurred with a similar incidence in the two groups. Those with a 4% or greater incidence were fever, viral hepatitis, and pneumonia. The incidence of sepsis, gastrointestinal hemorrhage, kidney failure, and convulsion was greater in the Int group. The incidence of abnormal liver function test results, hyperglycemia, headache, and abnormal kidney function was greater in the Rej group. Mean liver function test results decreased with time postrescue in both groups. Mean serum creatinine level increased from baseline to 18 months postrescue in both groups (1.44 to 1.51 mg/dL for Int patients, 1.14 to 1.48 mg/dL for Rej patients). We conclude tacrolimus is safe and effective rescue in liver transplant recipients with rejection or CyA intolerance.
Telomerase plays an important role in maintaining the stability of the chromosomes. Activity of telomerase has been detected in proliferating and immortalized cell lines and in a number of malignant tumors including invasive breast cancer. The aim of the study was to examine telomerase activity in ductal carcinoma in situ (DCIS), which is considered to be a precursor lesion of infiltrating breast carcinoma, using a PCR-based telomerase activity protocol (TRAP). We examined 35 samples obtained from histologically confirmed breast biopsies, including 13 normal breast tissues, 11 infiltrating ductal carcinoma (IDC), nine DCIS, and two DCIS with microinvasion. Telomerase activity was demonstrated in 8/9 samples of DCIS, both samples of DCIS with microinvasion, and all but one sample of IDC. Normal breast tissue had no demonstrable telomerase activity. Our results indicate that telomerase is activated frequently in early breast carcinogenesis, although its utilization as a biomarker in DCIS is questionable.
Explore the source record for details and available documents.
Methanococcus voltae harbors genetic information for two pairs of homologous [NiFe]-hydrogenases. Two of the enzymes contain selenocysteine, while the other two gene groups encode apparent isoenzymes that carry cysteinyl residues in the homologous positions. The genes coding for the selenium-free enzymes, frc and vhc, are expressed only under selenium limitation. They are transcribed out of a common intergenic region. A series of deletions made in the intergenic region localized a common negative regulatory element for the vhc and frc promoters as well as two activator elements that are specific for each of the two transcription units. Repeated sequences, partially overlapping the frc promoter, were also detected. Mutations in these repeated heptanucleotide sequences led to a weak induction of a reporter gene under the control of the frc promoters in the presence of selenium. This result suggests that the heptamer repeats contribute to the negative regulation of the frc transcription unit.
The time constant of the angular vestibulo-ocular reflex (aVOR), measured from the response to steps of rotation about a yaw axis, has frequently been estimated as a single exponential. However, the slow phase velocity envelope during per- or post-rotatory nystagmus is more accurately represented by two exponential modes. One represents activity in the vestibular nerve induced by deflection of the cupula, the other by activation that the input from the canals produces in the central velocity storage integrator. The sum of the cupula and the integrator responses describes the overall response of slow phase eye velocity and can be approximated by a double exponential. Frequently, there is a plateau in the initial portion of eye velocity response, but this may be masked by habituation, making the cupula contribution unobservable and impossible to estimate. Using a model-based technique to analyze responses with a clear plateau, we estimated peripheral and central vestibular time constants by double exponential fits to slow phase eye velocity. Cupular time constants were varied from 1 to 10 s to identify values that gave optimal fits of the data according to a Chi-square criterion. The mean cupular time constant for 10 human subjects was 4.2 +/- 0.6 s. Fits of the data were also good for time constants between 3.5 to 7 s, but not for 1 to 3 or 7.5 to 10 s. The estimated cupular time constants also fit responses where there was no plateau. In 8 monkeys, cupular time constants were estimated as 3.9 +/- 0.5 s, which agreed with those derived from activity in the vestibular nerve. There was no difference between monkey and human cupular time constants from these estimates. It is likely that the human cupular time constant is similar to that of the monkey and shorter than previously thought.
Platelet concentrate (PC) obtained from dogs with an automatic cell separator was stored in C4-cell separation sets with low gasdiffusionable Polyvinylchlorid (PVC) storage containers or in C4L-sets developed for storage with high gasdiffusionable Polyolefin(PO) containers, respectively. PC were stored for 10 days under permanent agitation at 22 degrees C (C4/22 degrees C, n = 10; C4L/22 degrees C, n = 11) or at 4 degrees C (C4L/4 degrees C, n = 6), respectively. Measurements were carried out directly after production of the PC, after 6 hours and then daily during the 10-day storage period. In the second part of this paper the results of pH, the concentration of bicarbonate, glucose, lactate and potassium ions as well as the activity of lactate dehydrogenase (LDH) are presented. The varying duration and intensity of the energy metabolism of the platelets and different part of glycolysis became obvious by the consumption of glucose and production of lactate, which differed significantly between the different storage conditions. Resulting from this, the mean pH decreased under the limit prescribed for human PC (pH = 6.3) already after a storage period of 3 days due to the slight capacity of gas diffusion in PVC-containers (C4/22 degrees C). In the PO-containers the pH fell below this limit at 22 degrees C (C4L/22 degrees C) after a storage period of 5 days and at 4 degrees C (C4L/4 degrees C) after 10 days. The latter reflects the high gas diffusion capacity of the PO-containers and the decreased metabolism activity at 4 degrees C. The increase of activity of LDH and of the concentration of potassium ions, which are localized in the cytosol of platelets, depended also on the different storage conditions and, thereby, reflected the different rapidity of increasing membrane permeability or the destruction of the cell membrane, respectively. The results of this study nearly are in agreement with the changes of platelet function shown in part I. Biochemical changes occur in canine platelet concentrates similar to those in human platelet concentrates during storage in dependency of the storage conditions, in part even with a higher rate or in a higher extent.
[NiFe]-hydrogenases generally carry the bimetallic Ni-Fe reaction center on their largest subunit. The [NiFeSe]-hydrogenase Vhu from Methanococcus voltae has an unusual subunit composition. Some of the amino acids participating in the formation of the reaction center are within a separate, very small subunit, called VhuU. It consists of only 25 amino acids and contains the selenocysteinyl residue, a ligand to the Ni atom. We have tested whether the special configuration of the Vhu-hydrogenase is of particular biochemical relevance. We have constructed a fusion subunit derived from the VhuA and VhuU subunits by generating a gene fusion which was inserted into the chromosome of M. voltae by gene replacement. The enzyme was purified and shown to be as active as the wild-type enzyme. M. voltae carries the genetic information for four different [NiFe]-hydrogenases. In addition to the Vhu-hydrogenase, a second selenium-containing enzyme, Fru, has been purified. Two selenium-free enzymes, Vhc and Frc, are homologues of Vhu and Fru, respectively. Their gene groups, vhc and frc are transcribed only upon selenium depletion. The selenium-containing subunit VhuU has been implicated in their negative regulation. However, cells containing the fusion hydrogenase still exhibited normal regulation of the vhc andfrc promoter activities as tested in reporter gene constructs. This indicates that the free VhuU polypeptide is not required for the negative regulation of the vhc or frc genes.
The similarity of the transcriptional apparatus of Archaea with that of Eucarya makes studies of their transcriptional regulation especially interesting. Such investigations are greatly facilitated by reporter genes. The concomitant analysis of several promoters for investigations of regulatory patterns requires different reporter genes. The archaeon Methanococcus voltae is a moderately halophilic mesophile. The treA gene from Bacillus subtilis appeared to be a good candidate for a reporter, since its product trehalase is salt-resistant. We show that it is indeed expressed under the control of a M. voltae promoter and that the enzyme is easily testable in cell lysates.
The immunomodulator AS101 has been previously shown to confer protection upon BALB/c mice infected with the intraerythrocytic parasite Babesia rodhaini (B. rodhaini). The present study focuses on the effect of AS101 administration on the acute phase of babesial infection where T helper cell subset patterns-TH1/TH2-were assessed in heavily infected mice. Secretion of cytokines of the TH1 subset (IL-2, IFN-gamma, IL-12) and of the TH2 subset (IL-10, IL-4) as well as TGF-beta was measured following the administration of AS101 2 weeks before parasite infection. Our results demonstrate that the parasites suppress IL-2 protein and IL-12 mRNA and that AS101 upregulates their secretion: IL-2, 8 u/ml vs 34 u/ml, respectively; IFN-gamma protein, 2370 pg/ml vs 4777 pg/ml, respectively. Conversely, babesial infection results in the upregulation of IL-10 and IL-4 proteins and TGF-beta transcripts, whereas AS101 downregulates their production: IL-10, 1800 pg/ml vs 360 pg/ml, respectively; IL-4, 58.3 pg/ml vs 24.5 pg/ml, respectively. A possible escape mechanism induced by B. rodhaini is suggested, starting with IL-10 inhibition of macrophage activities leading to a suppression of the TH1 response and of IL-2 in particular. It is therefore possible that AS101 may protect infected mice by activating cellular-mediated immunity and concurrently balancing the TH subset responses. It is suggested that AS101 may be effective as an antiparasitic drug.
BACKGROUND: Of the 20 known cytokeratins, CK-19 is expressed in normal urothelium, whereas the recently identified CK-20 is expressed in urothelial carcinoma cells but not in normal urothelial cells. The aim of this study was to examine whether CK-20 expression could serve as a noninvasive test in which malignant urothelial cells in urine are detected and monitored. METHODS: In the current study, the authors used reverse transcriptase-polymerase chain reaction (RT-PCR) methods to determine the expression of CK-20 in cells separated from the urine of patients with bladder carcinoma. Cells were obtained from the urine of 87 patients divided into the following 2 groups: 1) 14 healthy volunteers without any known history of transitional cell carcinoma (TCC), and 2) 73 patients with hematuria suspected for TCC of the bladder. For control purposes, CK-20 expression was examined in cells of 1) bladder carcinoma tumors of 5 patients, 2) blood of either patients with bladder carcinoma (n = 5) or healthy controls (n = 5), and 3) three different cell lines. RNA of the various cell pellets was extracted and RT-PCR was performed with CK-20 and CK-19 primers (CK-19 was used as a marker for normal epithelial cells). RESULTS: CK-20 amplification band (370 bp) was obtained with mRNA extracted from TCC cells of either bladder tumor or HT-29 line (a CK-20 colon carcinoma line). Sensitivity of the method was found to be 91%, whereas specificity was 67%. Among the 7 false-positive cases, 3 showed atypia, 3 hyperplasia, and 1 metaplasia, and 2 underwent previously successful TCC tumor removals, suggesting that the CK-20 test also responded to premalignant lesions. No false-positive cases were found in the healthy control group. No other preparation, including blood of the patients of with TCC, showed the CK-20 amplification band. CONCLUSIONS: These results indicate that CK-20 is a potential biomarker for noninvasive detection of bladder carcinoma by assaying uroepithelial cells from the voided urine specimen with RT-PCR.
This study utilized a non-experimental design to obtain information on a full range of domestic violence incidents brought before the Quincy, Massachusetts District Court, a model court. One limitation of previous research on spouse assaults using more sophisticated designs is that the target population has been restricted to specific subgroups of cases thereby limiting subsequent discussions of policy/practice implications of the findings vis-a-vis all spouse assault cases. To address this research "shortfall", we obtained permission from the Quincy District Court to examine all the spouse assault cases brought before the court during a 7-month period (June, 1995, through February, 1996). The findings show that in a full enforcement environment, victims took out restraining orders only against the most violent, criminally abusive men. Most men who were arrested for domestic violence had prior criminal histories for a variety of offenses. Domestic violence offenders appeared to be of two types: those with extensive and diverse criminal histories and those with little or no such involvement. However, active criminal justice intervention against domestic violence offenders appears to be primarily directed toward offenders already active in the criminal justice system.
OBJECTIVE: Cytokeratins are constituents of the intermediate filaments of epithelial cells which are expressed in various combinations depending on the epithelial type and the degree of differentiation. The recently identified cytokeratin-20 (CK-20) was found with immunohistochemical methods to be expressed in gastrointestinal epithelium, uroepithelial cells, and Merkel cells. Clues were also found for low expression of CK-20 in endometrial carcinoma cells. The aim of this study was to examine whether CK-20 expression can be measured in endometrial carcinoma with the more sensitive reverse transcriptase-polymerase chain reaction (RT-PCR) methods and therefore used as a potential diagnostic tumor biomarker for endometrial carcinoma. METHODS: In the present study we have used RT-PCR methods to determine expression of CK-20 in endometrial epithelial cells. Endometrial specimens were collected from 18 patients with endometrial carcinoma and 22 patients that underwent hysterectomy due to benign diseases. The specimens were prepared for both RT-PCR and immunohistochemical analysis. RNA, of the various cell pellets, was extracted and RT-PCR was performed with CK-20 and CK-19 primers (CK-19 was used as a marker for normal epithelial cells). Immunohistochemistry was carried out with the avidin-biotin-peroxidase complex method on formalin-fixed paraffin sections using CK-20 antibody. RESULTS: CK-20 amplification band (370 bp) was obtained with mRNA extracted from endometrial carcinoma cells of 17 of the patients with endometrial carcinoma (sensitivity, 94.4%). CK-20 was negative in 21 patients with benign endometrial disease (specificity, 91.3%). No positive results were obtained with the immunohistochemical methods. CONCLUSION: These results indicate that RT-PCR of CK-20, because of its high sensitivity, is a potential biomarker for detecting endometrial carcinoma.
Determination of Rb, Br, Se, Zn, Cu, Fe, and Br/Rb ratio in tissues of mice inoculated with colon and melanoma cancer cells is described. A group of 19 Balb/c mice inoculated with C26 colon carcinoma, 4 C57B1/6 mice inoculated with B16 melanoma, and 13 control mice of both kinds were under investigation. The study was conducted on samples of blood, liver, kidneys, colon, and skin, and the trace element levels in normal and inoculated mice were compared. The inoculation was by subcutaneous injection either at the back or intrafootpad. The blood samples were taken 1, 2, and 3 wk after inoculation, and after 4 wk all the animals were sacrificed. Two nondestructive, complementary analytical methods were used: a modified X-ray fluorescence (XRF) for solid tissue and particle-induced X-ray emission (PIXE) for blood samples. The detection limit (DL) in the PIXE method was 0.35 microg/g dry wt in 600 s counting time and in XRF, 1 microg/g dry sample for Rb, Br, Se and Zn and 2 microg/g for Cu and Fe in 200 s counting time. In all the cases studied, cancerous tissue developed at the site of the injection, and a significant difference in the trace element levels was observed between tissue samples obtained from normal and inoculated mice. The most pronounced effect was an increase in Rb level in the tumor by a factor ranging between 4 and 10 relative to normal tissue, with a corresponding decrease in the Br/Rb ratio (p < 0.05). Smaller changes were found in the Br, Se, Zn, and K levels. The changes in trace element levels in the inner organs were much smaller and seem to be influenced by the site of injection.