Electrochemical degradation of perfluoroalkyl and polyfluoroalkyl substances (PFASs) in groundwater.

Perfluoroalkyl and polyfluoroalkyl substances (PFASs) represent hazardous pollutants and are frequently detected in the environment, e.g. in contaminated groundwater. PFASs are persistent to biodegradation and conventional oxidation processes such as ozonation. In this study electrochemical degradation of PFASs on boron-doped diamond (BDD) electrodes is demonstrated. Experiments were performed with model solutions and contaminated groundwater with a dissolved organic carbon (DOC) content of 13 mg/L. The perfluorinated carboxylic acids (PFCAs) perfluorobutanoate, perfluoropentanoate, perfluorohexanoate, perfluoroheptanoate and perfluorooctanoate, and the perfluorinated sulfonic acids (PFSAs) perfluorobutane sulfonate, perfluorohexane sulfonate, perfluorooctane sulfonate and 6:2 fluorotelomer sulfonate were detected in the groundwater samples. At PFAS concentrations ranging from 0.26 to 34 mg/L (0.7 to 79 µM), the degradation of PFASs was achieved despite of the high DOC background. Pseudo first-order kinetic constants of PFSA degradation increased with the increase of carbon chain length. Fluoride formation as well as the generation of PFCAs with shortened chain lengths was observed. Inorganic byproducts such as perchlorate were also formed and have to be considered in further process optimization.

Effects of electric polarization of indium tin oxide (ITO) and polypyrrole on biofilm formation.

The influence of electric polarization on primary adhesion and on biofilm formation was investigated. As substrata, indium tin oxide (ITO) and polypyrrole coatings were used because of their electric conductivity. The materials were polarized from -600 mV to +600 mV, switching every 60 seconds. Control was non-polarized substrata. Primary adhesion under this regime was not strongly influenced, however, the morphology of the primary biofilm was obviously different from that of the control. Biofilm formation of the natural population of non-chlorinated drinking water, supplemented with nutrient in low concentration, was determined over 164 hours. While the biofilm on the control surface developed to a thickness of about 100 microm, on the pulsed polarized surface it reproducibly developed only to a very thin biofilm. Faster switching of the polarization (10 second) had no further influence. If the polarization routine was reduced to only twice a day (one hour), no influence on biofilm development was observed. These results indicate that fluctuating polarization at a rate of once per minute inhibits the physiological processes during biofilm formation during one week. Investigations are in process to determine further details of this effect in order to employ it for inhibition of biofouling.

Interobserver reproducibility of Gleason grading of prostatic carcinoma: urologic pathologists.

Gleason grading is now the most widely used grading system for prostatic carcinoma in the United States. However, there are only a few studies of the interobserver reproducibility of this system, and no extensive study of interobserver reproducibility among a large number of experienced urologic pathologists exists. Forty-six needle biopsies containing prostatic carcinoma were assigned Gleason scores by 10 urologic pathologists. The overall weighted kappa coefficient kappa(w) for Gleason score for each of the urologic pathologists compared with each of the remaining urologic pathologists ranged from 0.56 to 0.70, all but one being at least 0.60 (substantial agreement). The overall kappa coefficient kappa for each pathologist compared with the others for Gleason score groups 2-4, 5-6, 7, and 8-10 ranged from 0.47 to 0.64 (moderate-substantial agreement), only one less than 0.50. At least 70% of the urologic pathologists agreed on the Gleason grade group (2-4, 5-6, 7, 8-10) in 38 ("consensus" cases) of the 46 cases. The 8 "nonconsensus" cases included low-grade tumors, tumors with small cribriform proliferations, and tumors whose histology was on the border between Gleason patterns. Interobserver reproducibility of Gleason grading among urologic pathologists is in an acceptable range.

Interobserver reproducibility of Gleason grading of prostatic carcinoma: general pathologist.

Only a few large studies of interobserver reproducibility of Gleason grading of prostatic carcinoma exist. Thirty-eight biopsies containing prostate cancer were distributed for Gleason grading to 41 general pathologists in Georgia. These cases had "consensus" Gleason grade groups (2-4, 5-6, 7, and 8-10) that were agreed on by at least 7 of 10 urologic pathologists. The overall kappa (kappa) coefficient for interobserver agreement for these 38 cases was 0.435, barely moderate agreement, with a kappa range from 0.00 to 0.88. There was consistent undergrading of Gleason scores 5-6 (47%), 7 (47%) and, to a lesser extent, 8-10 (25%). In cases with consensus primary patterns, there was consistent undergrading of patterns 2 (32%), 3 (39%), and 5 (30%). Pattern 2 was often (17%) mistaken for pattern 3. Pattern 4 was often undergraded (21%) and also mistaken for pattern 5 (17%). The most significant (P < .005) demographic factor associated with better interobserver agreement was having learned Gleason grading at a meeting or course. We believe that Gleason grading can be learned to a satisfactory level of interobserver reproducibility and have undertaken additional studies that support this belief.

Alterations of adenomatous polyposis Coli (APC) gene in oral squamous cell carcinoma.

The present study assessed the roles of Adenomatous Polyposis Coli (APC) tumor suppressor gene during oral carcinogenesis. Reduction of APC transcript levels and APC loss of heterozygosity (LOH) were found in 39% (7/18) and 29% (10/34) cases of oral squamous cell carcinoma (OSCC), respectively. The apparent APC heterozygosity (27%) in non-cancerous matched oral tissue (NCMOT) adjacent to OSCC at an exon 11 locus was significantly lower than normally found in the Taiwanese population (49%). These findings suggest that the allelic status of APC could indicate a cancer risk. No polymorphism of 11307 allele of APC was identified in NCMOT or OSCC. Our data indicated that alterations of APC are frequent molecular changes of OSCC. Advances in understanding of the APC alterations that accompany OSCC development might provide a means for early diagnosis and possibly new therapeutic strategies.

Reference electrodes based on conducting polymers.

Several attempts to produce conducting polymer based all-solid-state reference electrodes are presented. Open circuit potential of conducting polymers is redox sensitive and Donan equilibrium dependent. Therefore, more sophisticated constructions are necessary. Most promising were bilayers composed of conducting polymers with different ion-exchanger properties.

Needle biopsies of the prostate: what constitutes adequate histologic sampling?

OBJECTIVE: The automated biopsy gun and increased screening for adenocarcinoma of the prostate have led to increased numbers of biopsies with only tiny foci of prostatic carcinoma. Consequently, the risk of failing to sample a small focus of carcinoma histologically has increased as well. Most pathologists routinely sample prostatic needle biopsies at more than 1 level. An expert panel has recently suggested that prostatic needle biopsies be sampled at at least 2 levels. However, there have been no studies measuring the amount of additional tissue sampled by multiple levels versus 1 level. METHODS: Forty-two prostatic needle biopsies were serially sectioned at 4-microm levels. Hematoxylin-eosin-stained slides were prepared from every fifth section. The total length of each biopsy was compared with the length sampled by 1 level (50% through the block) and 3 levels (25%, 50%, and 75% through the block). RESULTS: Sampling the tissue at 1 level missed an average of 23.4% of the total biopsy length. Sampling the tissue at 3 levels significantly improved this average to 7% (P = .0001). CONCLUSIONS: This study shows that a single histologic section of a prostatic needle biopsy often fails to sample a significant portion of available tissue. This could occasionally result in failure to sample a small focus of prostatic carcinoma. The authors recommend that prostatic needle biopsies be routinely sampled at 3 levels (approximately 25%, 50%, and 75% through the block).

p16 (CDKN2/MTS1) gene deletions are rare in prostatic carcinomas in the United States and Japan.

PURPOSE: The incidence of clinically apparent prostatic carcinoma is much higher in the United States than in Japan. Alterations in the p16 tumor suppressor gene have been identified in various tumor types, including cultured prostatic carcinoma cell lines. We studied the possible deletions of either exon 2 or 3 of this gene in primary clinical prostatic carcinomas from Japan and the United States. MATERIALS AND METHODS: Genomic DNA was extracted from 36 formalin-fixed, paraffin-embedded clinical prostatic carcinomas from Japan and 27 carcinomas from the United States. Exons 2 and 3 of the p16 gene were amplified using comparative multiplex polymerase chain reactions (PCR) and then analyzed for possible deletions of either exon. RESULTS: Two out of 36 (5.6%) carcinomas from Japan clearly demonstrated deletion of p16 exon 2, but this deletion was not detected in any of the 27 carcinomas from the United States. CONCLUSIONS: Although slightly higher in Japan than in the United States, the frequency of p16 exon deletions in clinical prostatic carcinomas is very low, and probably is not important in the development of this neoplasm.

Extensive extramedullary disease in myeloma. An uncommon variant with features of poor prognosis and dedifferentiation.

BACKGROUND: The development of extramedullary plasmacytomas and elevated serum lactic dehydrogenase (LDH) in myeloma indicates poor prognosis. A 75-year-old man was diagnosed with immunoglobulin (Ig) A, lambda myeloma when he developed pathologic rib fractures, hypercalcemia, and anemia. After 6 months of treatment with melphalan and prednisone, he was in complete remission as evidenced by the disappearance of the monoclonal protein in the serum and free light chain in the urine. Eight months after diagnosis, his disease took an unusual course with the simultaneous development of plasmacytomas in the skin, breast, stomach, and pancreatic head, complicated by severe upper gastrointestinal bleeding and obstructive jaundice. METHODS: Immunohistochemical staining of the marrow and breast mass was done using monoclonal antibodies against B-cell and T-cell antigens as well as kappa and lambda light chains. In situ hybridization was performed to detect ras oncogene overexpression in the breast mass. RESULTS: Immunohistochemical staining of the original marrow and breast mass was positive for IgA and lambda, confirming the identical clonal origin of the plasma cells. The disorder expressed elevated serum LDH, both at diagnosis and relapses. Features of dedifferentiation were expressed by the disappearance of myeloma protein in the serum at relapse, absence of marrow plasma cell infiltration, and development of multiple extramedullary plasmacytomas. There was no overexpression of H-ras or N-ras oncogenes by in situ hybridization of the plasmacytoma from the breast. The patient died shortly after the development of the extramedullary plasmacytomas. CONCLUSIONS: The simultaneous appearance of plasmacytomas in multiple extramedullary sites heralds a change of clinical behavior in myeloma. When accompanied by the disappearance of serum myeloma protein, and marrow plasma cell infiltration, and serum LDH elevation, the disorder may follow a fulminant course.

Multiple genetic alterations in hamster pancreatic ductal adenocarcinomas.

Pancreatic ductal adenocarcinomas induced in the Syrian golden hamster (SGH) by N-nitrosobis(2-oxopropyl)amine share many similarities with the human disease, including mutations of the K-ras oncogene. In vitro carcinogenesis studies with immortal SGH pancreatic duct cells indicate that neoplastic transformation in this system can occur without mutational inactivation of p53 suppressor gene. In this study we extend the genetic analysis of the in vivo SGH model to increase the number of cases analyzed for the status of K-ras and to determine further the spectrum of alterations involved; we have studied the status of the p53, DCC, and Rb-1 suppressor genes and the status of the mdm2 oncogene, which can involve p53 indirectly. The partial SGH-coding sequence of mdm2 and DCC was determined. K-ras mutation in the second position of codon 12 was present in 17 of 19 (90%) of tumors. Immunohistochemistry and single strand conformation polymorphism analysis showed no evidence of p53 mutation in 21 tumors. RNase protection assays showed overexpression of mdm2 in 5 of 19 (26%) tumors. Semiquantitative reverse transcription-PCR analysis showed a complete or partial loss of DCC expression in 10 of 19 (53%) neoplasms and of Rb-1 (42%) expression in 8 of 19 tumors when compared to matched controls. Deregulation of these genes appears to be significant in SGH pancreatic carcinogenesis as indicated by their frequencies. However, the fact that 6 tumors showed either only a K-ras mutation or the absence of alterations of the 5 genes analyzed indicates that additional as yet unstudied or unknown genes are also involved in SGH pancreatic duct carcinogenesis.

In vitro carcinogenesis of hamster pancreatic duct cells: cellular and molecular alterations.

Neoplastic transformation of Syrian golden hamster (SGH) pancreatic duct cells was induced by in vitro treatment with the direct-acting carcinogens N-methylnitrosourea (MNU) and N-(2-hydroxypropyl)nitrosourea (HPNU), with subsequent selection by sustained culture in serum- and epidermal growth factor (EGF)-deprived medium. The present study examines the efficacy of serum and EGF deprivation as a selection pressure and the effect of the carcinogen dose, frequency and interval of exposure on tumorigenesis and K-ras mutation. Selection of carcinogen-initiated duct cells by serum and EGF deprivation is highly reproducible and effective, increasing the incidence of tumors from 26 to 93% for MNU or from 0 to 100% for HPNU. SGH pancreatic duct cells exposed to 0.5 mM MNU for 13 weeks (long-treatment schedule) produced K-ras mutations at codon 12 in six of six tumors. However, when cells were exposed to 0.125, 0.25 or 0.5 mM MNU daily for 5 days (short-treatment schedule), mutations of K-ras at codon 13 were identified in four of 16 tumors, the remaining 12 showing no mutations. Duct cells exposed to 0.5 mM HPNU by the short-treatment schedule produced K-ras mutations in codon 13 in six of six tumors, as contrasted to 12 tumors that developed from cells exposed to 0.125 or 0.25 mM HPNU, which all contained K-ras codon 12 mutations. The current experiments demonstrate that K-ras mutation in pancreatic carcinogenesis in vitro by MNU or HPNU can be modified by the nature and dose of the carcinogen as well as the frequency and duration of exposure.

Genomic p53 mutation in a chemically induced hamster pancreatic ductal adenocarcinoma.

This study investigates possible alterations in exons 5 through 8 of the p53 gene and altered p53 protein expression in the Syrian golden hamster model of pancreatic ductal adenocarcinoma. In the Syrian hamster p53 sequence, 1469 base pairs are presented for the genomic regions surrounding exons 5 through 8, along with the primer sequences specific for the enzymatic amplification of the individual exons. Single-stranded conformation polymorphism was analyzed on the products obtained by enzymatic amplification of hamster genomic DNA extracted from 2 transplantable pancreatic ductal adenocarcinomas, 6 groups of N-methylnitrosourea (MNU)-treated pancreatic duct cells, and 17 MNU-induced pancreatic ductal adenocarcinomas. The two transplantable adenocarcinomas were a well-differentiated ductal adenocarcinoma established from a N-nitrosobis(2-oxopropyl)amine-induced tumor and a poorly differentiated ductal adenocarcinoma established from a spontaneous tumor. An altered mobility indicated a conformational change in the first part of exon 5 in the solid form of the well-differentiated ductal adenocarcinoma. Direct sequencing of the amplified product revealed an A-->C transversion in codon 135, which corresponds to codon 132 in the human p53 gene. A conformational change in exon 7 was observed in 1 of 6 MNU-treated cell samples, and none of the 17 resultant tumors. Direct sequencing confirmed a deletion of one C of the three in codon 263, which generates a frameshift mutation. No conformational change was observed in any other products. Positive staining with PAb240 or DO7 antibodies against human p53 or with an antibody generated in our laboratory against the hamster p53 fusion protein was observed only in the solid form of well-differentiated ductal adenocarcinoma and in rare cells scattered in 4 of 28 MNU-induced tumors analyzed. This study provides a system to analyze p53 gene alterations in the hamster and is the initial report of a specific p53 mutation in a hamster pancreatic ductal adenocarcinoma.

K-ras mutation is an early event in pancreatic duct carcinogenesis in the Syrian golden hamster.

K-ras oncogene mutation has been shown to be a frequent event in pancreatic ductal adenocarcinomas induced by the carcinogen N-nitroso-bis(2-oxopropyl)amine in the hamster. The present study examines the mutational status of the K-ras oncogene in lesions that precede the appearance of invasive ductal adenocarcinomas. Syrian golden hamsters (80-100 g) received 12 weekly doses of 15 mg/kg N-nitroso-bis(2-oxopropyl)amine and were serially sacrificed at 8, 12, 14, 16, or 24 weeks following the initiation of treatment. Ten microns-thick sections of formalin-fixed paraffin-embedded pancreas were examined for hyperplasia, papillary hyperplasia, carcinoma in situ, and invasive and metastatic ductal carcinoma. Marked lesions of interest were scraped from the slide, subjected to polymerase chain reaction-mediated amplification of the first exon of the K-ras gene, and probed by oligonucleotide-specific hybridization for mutations at codon either 12 or 13. Of 186 samples assayed, K-ras codon 12 mutations were detected in 26% of hyperplasias, 46% of papillary hyperplasias, 76% of carcinoma in situ, 80% of adenocarcinomas, and 43% of lymph node metastases. Codon 12 mutations were exclusively G to A changes at the second position. Codon 13 mutations were only detected in 9 of 168 samples. These results suggest that K-ras activation is an early event in N-nitroso-bis(2-oxopropyl)amine-induced pancreatic carcinogenesis in the hamster.

Analysis of ras gene mutations in rainbow trout liver tumors initiated by aflatoxin B1.

The suspect human hepatocarcinogen aflatoxin B1 (AFB1) is a well-known potent initiator of hepatic tumors in rainbow trout (Oncorhynchus mykiss). Both hepatocellular carcinomas and mixed hepatocellular/cholangiocellular carcinomas are induced by AFB1 in trout, with the mixed form predominating. We previously isolated two c-ras genes from trout liver cDNA, and in the present study we analyzed DNA from 14 AFB1-induced trout liver tumors for point mutations in exon 1 of both genes. Using the polymerase chain reaction (PCR) and oligonucleotide hybridization methods, a high proportion (10/14) of the AFB1-initiated tumor DNAs showed evidence of activating point mutations in the trout c-Ki-ras gene. Of the 10 mutant ras genotypes, seven were codon 12 GGA----GTA transversions, two were codon 13 GGT----GTT transversions, and one was codon 12 GGA----AGA transition. Nucleotide sequence analysis of cloned PCR products from four of these tumor DNAs provided definitive evidence for two codon 12 GGA----GTA mutations, one codon 12 GGA----AGA mutation, and one codon 13 GGT----GTT mutation, in complete agreement with the oligonucleotide hybridization results. No mutations were detected in exon 1 of a second trout ras gene also expressed in liver, nor in DNA from control livers. This is the first report of experimentally induced ras gene point mutations in a lower vertebrate fish model. The results indicate that the hepatocarcinogen AFB1 induces c-Ki-ras gene mutations in trout similar to those in rat liver tumors.

Expression of ras genes in rainbow trout liver.

Members of the ras gene family have been studied in a variety of species. Two ras genes expressed in the normal liver of rainbow trout, ras-1 and ras-2, as well as a portion of a genomic ras-1 allele, are described for the first time in this report. Over 500 bp of trout ras-1 and at least 300 bp of trout ras-2 genes expressed in normal liver have been sequenced; DNA homology to the human ras genes ranges from 76.8% to 87.1%. The base changes resulting from over 400 million years of evolutionary divergence between the species were primarily silent, with few changes in protein sequence. The partial DNA sequence of the genomic ras-1 allele has 86.8% homology to the first two exons of human c-Ha-ras, and its intron has several conserved sequences characteristic of vertebrate intron-exon junctions. However, the predicted amino acid sequence of trout ras-1 differs at only one of the first 172 amino acid residues from human c-Ki-ras with the alternate exon 4b. Since trout ras-1 differs at 17 and 18 residues from the human c-N-ras and c-Ha-ras proteins, respectively, over this region, we conclude that trout ras-1 is a c-Ki-ras gene. The highly conserved nature of this gene suggests that the ras p21 protein has identical functions in normal and neoplastic cells among higher and lower vertebrates.

Activation of K-ras in transplantable pancreatic ductal adenocarcinomas of Syrian golden hamsters.

High mol. wt genomic DNA was prepared from normal hamster pancreas and the solid and ascites variants of two different hamster transplantable carcinomas, one induced by N-nitrosobis(2-oxopropyl)amine and the other spontaneously occurring. This DNA was transfected into NIH/3T3 cells and resulted in cells that were capable of forming tumors when injected into nude mice. Analysis of the nude mouse tumors by Southern blotting revealed the presence of a band specific for hamster K-ras. Polymerase chain reaction (PCR)-mediated amplification of the K-ras codon 12-13 region of genomic DNA prepared from the transplantable tumors produced a 117 bp fragment which was analyzed by both allele-specific oligonucleotide hybridization and direct DNA sequencing. Oligonucleotide hybridization with probes specific for changes in the first or second position of codons 12 or 13 detected a G to A transversion in the second position of codon 12 in the chemically induced transplantable tumor, and a G to A change in the second position of codon 13 in the spontaneously occurring transplantable carcinomas. The result obtained for the chemically induced tumor was confirmed by direct dideoxy sequencing of the PCR-amplified product. These findings are the first to show a specific oncogene activation in an experimental pancreatic tumor model and also parallel the results recently reported for K-ras mutations in human pancreatic carcinoma.

The effects of extracts from neurosecretory cells in the anterior vena cava and pharyngo-ophthalmic vein upon the hearts of intact free-moving octopuses.

Recordings of pressure and frequency were made from the hearts of free-moving Octopus vulgaris. The effects of extracts from neurosecretory endings in the anterior vena cava (AVC) and the pharyngo-ophthalmic vein (POV), injected through fine cannulae into a branchial heart, efferent branchial vessel or the dorsal aorta, were studied and compared with the effects of acetylcholine, 5-hydroxytryptamine, adrenaline, histamine and tyramine. AVC and POV extracts each produce a different spectrum of effects, unlike those of any of the drugs tested. AVC extract is effective at doses of less than 2% of the material extractable from a single vein per kg, increasing the force and amplitude of the heartbeats. With a natural release point just upstream of the branchial hearts the AVC material must be relevant to the normal performance of the hearts. POV extract is effective only at doses equivalent to several veins per kg, and is unlikely to have a role in cardiac regulation. Section of the visceral nerves did not affect the action of drugs or extracts, indicating that effects were not indirectly mediated via the CNA. Further experiments were made with hearts and the aorta in vitro with effects that did not always parallel those found in vivo. Reasons for these differences are discussed.