Inverse relationship between microsatellite instability and K-ras and p53 gene alterations in colon cancer.

Some studies have shown an inverse relationship between microsatellite instability in colon cancer and mutations in p53 and K-ras, whereas others have not. We therefore evaluated these features in a population-based sample of 496 individuals with colon cancer. Microsatellite instability was determined by a panel of 10 tetranucleotide repeats, the Bethesda consensus panel of mono- and dinucleotide repeats, and coding mononucleotide repeats in transforming growth factor-beta receptor type II, hMSH3, BAX, hMSH6, and insulin-like growth factor receptor type II. Mutations in codons 12 and 13 in K-ras were evaluated by sequencing. p53 overexpression (as detected by immunohistochemistry) was used as an indicator of p53 mutation; this was evaluated in 275 of the tumors. K-ras mutations were present in 33.2% of tumors, p53 overexpression in 51.5%, and microsatellite instability (as determined by the Bethesda consensus panel) in 12.5%. K-ras mutations were significantly less common in unstable tumors than stable tumors (11.8% versus 36.9%, P: < 0.001). p53 overexpression was significantly less common in unstable tumors than stable tumors (20.0% versus 55.7%, P: < 0.001). These inverse relationships between microsatellite instability and ras gene mutations and p53 overexpression were shown to be independent of tumor site in logistic regression analyses. All other measures of instability also showed statistically significant inverse relationships independent of tumor site with alterations in ras and p53, and instability results determined by the panel of 10 tetranucleotide repeats were highly significantly related to those determined by the Bethesda consensus panel. Coding mononucleotide repeat mutations were significantly more common in unstable tumors than stable tumors (85.7% versus 1.0%, P: < 0.001). We conclude that there is an inverse relationship between microsatellite instability and mutations in p53 and K-ras, and that the molecular profile of colon cancers with microsatellite instability is characterized by relatively infrequent mutations in K-ras and p53 and relatively frequent mutations in coding mononucleotide repeats.

Flexible and insulative Plasmalene wire coatings for biomedical applications.

A novel polymeric material, Plasmalene, was synthesized and tested as a functional hermetic polymer coating on Gold, Platinum, and MP35N wire substrates. The flexible Plasmalene polymer coatings were synthesized onto wire substrates by vacuum processes which included radio frequency (RF) initiated plasma polymerization and thermal vapor deposition of methane and xylylene monomers. Mechanical and electrical tests were performed on 75 micron wires coated with either Plasmalene, Parylene, or Teflon polymers. The coated wire substrates were mounted on a flex testing machine that was submerged in a saline bath. A +5VDC electrical bias was continuously applied between the wire substrates and the bath. The flex testing machine mechanically stressed the coatings for 50,000 cycles with tensile and compressive loadings of up to 11% strain. The cyclic strain was applied through a total flexural arc of 170 degrees. Electrical leakage currents were measured through the polymer films to the saline bath at +5VDC versus a Ag/AgCl reference electrode. Analysis of the leakage current data for the Parylene and Teflon coated wires indicated that there was a significant (alpha = 0.05) increase, up to microampere levels, in leakage currents through the films as a result of repeated cyclic stressing. Plasmalene coated wires showed no significant increase in initial picoampere leakage currents with cycling and therefore, demonstrated that a superior insulative and mechanically durable film can be synthesized as a wire encapsulant.

Cold- and ethanol-induced hypothermia reduces cellular levels of mRNA-encoding Thyrotropin-Releasing Hormone (TRH) in neurons of the preoptic area.

Thyrotropin-releasing hormone (TRH)-containing neurons have been implicated in the central control of body temperature. TRH-containing neurons are located in brain areas known to influence body temperature, and TRH injected into these areas can produce changes in body temperature. While these lines of evidence support the view that central TRH is involved in thermoregulation, it has been difficult to confirm that TRH-containing neurons of the preoptic area are involved in this process. We used a different approach to test this hypothesis, based on recent evidence that changes in cellular levels of neuropeptide mRNA are linked to changes in neurosecretory processes. Hence, we predicted that if TRH neurons of the preoptic area are involved in body temperature regulation, cellular levels of TRH mRNA would be altered in animals in which body temperature had been experimentally altered. TRH mRNA levels were measured by in situ hybridization histochemistry in neurons of the preoptic area (POA) of animals that had been exposed to cold (5 degrees C) or that had been given a hypothermic dose of ethanol. Cellular levels of TRH mRNA were reduced by both treatments. However, cellular levels of the mRNA-encoding gastrin-releasing peptide were not affected by these treatments in neurons of the POA, indicating that hypothermia exerted selective effects on TRH neurons in this brain region. Considering that both cold exposure and ethanol administration increase blood pressure, that the POA contains neurons which are both thermosensitive and barosensitive, and that TRH has been implicated in the control of blood pressure, we manipulated arterial blood pressure pharmacologically without changing body temperature to determine whether TRH neurons were also responsive to cardiovascular changes. Infusions with either nitroprusside, a vasodilator, or phenylephrine, a vasoconstrictor, produced significant changes in arterial blood pressure and heart rate, but did not affect TRH mRNA in the POA. These findings demonstrate that TRH neurons of the POA are thermoresponsive, supporting the view that they play a role in the central control of body temperature.

Functional hermetic encapsulation of integrated circuits.

This study demonstrates a new process technology for synthesizing thin films which should be applicable to a variety of implantable devices of different substrate composition and geometries. This new technology, termed the plasma polymerization/vapor deposition (PP/VD) technology, combines the features of surface cleaning, surface treatment, and hermetic encapsulation into a series of in situ vacuum processes. This PP/VD process has produced functional hermetic encapsulating polymer films with a total thickness of 5 microns, independent of the composition, dimensions or geometry of the substrates. CMOS integrated circuits coated by this process and immersed in Ringer's solution and tested for a year, demonstrated unchanging device performance parameters characteristic of hermetically encapsulated integrated circuits.

The influence of Encephalitozoon cuniculi on neural tissue responses to implanted biomaterials in the rabbit.

Laboratory rabbits are commonly used for testing the tissue response of neural device biomaterials. Rabbits of many colonies in the U.S. are infected by the intracellular microsporidian parasite, Encephalitozoon cuniculi, with rates of infection ranging from 15 to 76% (1). Several authors have suggested that infection by this parasite may alter immune system response and experimental results. We report that infection by E. cuniculi made the interpretation of results more difficult and altered the animals' responsiveness to implanted platinum wires coated with various polymers such as glow discharge methane, Parylene C, or polyimide. Edema, neuronal and glial reaction, and inflammatory responses to the coated wires were quantitated at four sites in each animal. Inconsistency of response in all measured parameters was found, both between animals and between sites in infected animals. Infected animals showed the greatest variability, primarily in the degree of inflammatory reaction. Parylene C was found to induce the most severe inflammatory reaction, an unexpected finding. No consistent reaction to any of the coating materials was found in this study. We believe that this variability in response was primarily due to infection by E. cuniculi. Our results suggest that rabbits should not be used for tissue compatibility testing of neural device biomaterials until the animals are free of E. cuniculi infestation as demonstrated by serologic screening.

Electrical insulation of implantable devices by composite polymer coatings.

Protection of implanted integrated circuits has required hermetic sealing, usually in metal containers and usually in package sizes that would preclude implantation in small and confined areas of the body. We have developed a method whereby ultrathin (10 micron) composite films consisting of glow discharge and vapor deposited polymers can be placed directly over integrated circuit substrates to provide protection from water and ions for up to 30 days (our present test limits). Our paper describes the reactor, surface preparation, and polymerization conditions necessary to obtain the water/ion resistant coatings. Results indicate little change in leakage current when comb patterns with 10 micron line widths and our insulating composite coatings are exposed to physiological saline solution and a 3 VDC bias.

The adhesion of glow--discharge polymers, Silastic and Parylene to implantable platinum electrodes: results of tensile pull tests after exposure to isotonic sodium chloride.

Thin organic coatings commonly are used for insulating microelectrodes and electronic packages designed for implant applications. The adherence of these coatings to the underlying substrates is a key parameter in their selection for various devices. Instron pull tests were performed on glow-discharge polymerized monomers, Parylene-N, medical-grade Silastic and various epoxies. The application of a thin coating of glow-discharge polymerized methane under a thicker Parylene-N coating improved the adhesion of the latter to the underlying substrate in isotonic sodium chloride solution and during accelerated testing conditions done by boiling.

Physiologic features of the canine esophagus: effects of tranquilization on esophageal motility.

Esophageal motility was studied in clinically normal dogs. Mean values as to the time and pressure functions of esophageal peristaltic pressure curves are presented. Effects of two dosage levels of tranquilization on the time and pressure functions are reported.

Bioevaluation of plasma polymerized films in skeletal muscle.

Plasma polymerized ethylene (PPE), styrene (PPS), and chlorotrifluoroethylene (PPCTFE) were synthesized by exposing the monomeric gases to an inductively coupled radio frequency "glow-discharge" field. The polymer films were deposited on poly(dimethyl) siloxane (medical grade Silastic), which was then surgically implanted in rat paravertebral muscle for periods up to 84 weeks. The biocompatibility of the plasma deposited films and uncoated Silastic was evaluated by qualitative (graded inflammatory cell response) and quantitative (connnective tissue capsule thickness) techniques as a function of time. The morphological features of the connective tissue capsule and the plasma polymerized films were examined by SEM after 75 weeks of implantation. Results showed that the acute inflammatory cell migration around PPS and PPCTFE was at a maximum in 2 weeks, decaying to control levels in 4 to 8 weeks. The PPE response was judged as less than the control response up to 4 weeks. After 8 weeks no qualitative difference could be detected between the plasma polymerized films and Silastic. On the other hand, a quantifiable change in fibrous capsule response as a function of time and material was noted until 24 weeks. From these data we conclude that these types of films do not elicit an untoward foreign body reaction at a skeletal muscle implant site in rats.

Performance of implanted biogalvanic pacemakers.

Seventeen unipolar cardiac pacemakers powered by hybrid biogalvanic cells were implanted in dogs. Long term clinical effects and electrical performance in vivo of the generators were investigated. The biogalvanic cells were designed to provide 50 years of generator operation. No systemic pathological changes which would be attributed to the generator or the biogalvanic cell were observed. Local reaction to implants was mild and not significantly different from the one observed in conventional pacemaker implantations to date. The electrical performance of the biogalvanic cells was very encouraging. Final steady-state cell voltage levels of .65V to .75V were reached in 100--150 days after implantation and remained constant in 3 units to date which represents over 36 months after implantation for each generator.