An in vitro study of the effect of the optimal irrigation solution conditions during canine articular surgery.

OBJECTIVES: To establish whether chondrocyte viability, matrix degradation and the induction of proteolytic gene expression in canine cartilage is independent of irrigation fluid osmolality and time following exposure to the irrigation fluid. METHODS: Canine cartilage explants were exposed to one of three different solution types i) Culture medium (270-280 mOsmol/kg) ii) NaCl 0.9% (302 mOsmol/kg) iii) NaCl 0.9% with sucrose (600 mOsmol/kg). Chondrocyte viability and selected proteolytic gene expression were measured at two time points; immediately following exposure and 24 h following exposure. The media samples at 24 h following exposure were assessed for sulphated glycosaminoglycan (sGAG) release. RESULTS: In all samples, no cell death was observed across the superficial or deeper layers of the cartilage. When adjusting for time, gene expression was not shown to be dependent on solution type. However for all solution types, Matrix Metalloproteinase 13 (MMP13) and A Disintegrin and Metalloproteinase with Thrombospondin Motifs 5 (ADAMTS5) expression was significantly decreased in cartilage samples at 24 h post exposure comparatively to samples tested immediately post exposure. No significant differences were identified in the relative sGAG release between the solution types. CLINICAL SIGNIFCANCE: Arthroscopic solution irrigation of cartilage explants had no effect on cell viability or proteinase production. At present there is no indication to optimise irrigation fluid osmolarity, as conventional arthroscopic solution was not deleterious to healthy cartilage in this model.

Avoiding state intervention in not-for-profit/for-profit affiliations.

States attorneys general recently have intervened in transactions involving not-for-profit organizations that wish to convert to or transfer assets to for-profit status. These interventions are significant for two reasons: first, they reflect a tendency amount state attorneys general to involve states in the governance of not-for-profit healthcare organizations, and second, they demonstrate that attorneys general are paying renewed attention to the legal obligation of such organizations to provide a community health benefit, which imposes a charitable trust on not-for-profit healthcare organizations and the fiduciary duties of care and loyalty to that charitable trust on the organizations' officers and directors. To avoid state intervention in such transactions, officers and directors of not-for-profit organizations need to understand the circumstances under which attorneys general justify such intervention, which include the undervaluation of the organization's charitable assets; lack of a private letter ruling from the IRS; failure to adequately consider alternatives to the transaction; conflicts with the best interest of the organization; and inadequate responses to the attorney general's requests for information.

Melanogenesis-targeted anti-melanoma pro-drug development: effect of side-chain variations on the cytotoxicity of tyrosinase-generated ortho-quinones in a model screening system.

A set of 26 substituted phenols, 10 of which were synthesised in our laboratories, were tested for their rate of oxidation by mushroom tyrosinase in vitro as determined by oximetry and spectrophotometry and for their cytotoxic action in a model system. With one exception (4-hydroxybenzoic acid) all the agents tested were oxidised to the corresponding ortho-quinones. The maximum rates of oxidation varied between 15.1 +/- 0.59 nmoles oxygen consumed per minute (4-(2-thioethylthio)phenol) and 372.9 +/- 5.61 nmoles O2/ min. (4-(2-Hydroxyethylthio)phenol) in a reaction system comprising 300 units tyrosinase and 200 microM substrate. The rates of generation of quinone were in close agreement with these oximetric data. Some anomalies in oxygen stoichiometry were observed due to reoxidation of reaction products. Four categories of compounds were tested: those known to undergo side-chain cyclisation (such as tyrosine) (Group A), alkylphenols of increasing chain length with or without terminal hydroxyl groups (Group B), compounds with charged or bulky side-chains (Group C) and agents with oxy-, thio- and selenyl-ether side-chains (Groups D, E and F). In the majority of cases, the cytotoxicity, measured by the reduction of thymidine incorporation in cells exposed for 30 min to the agent in the presence of tyrosinase, reflected the rate of oxidation and is ascribed to the toxic action of the derived ortho-quinone. Tyrosinase-dependent cytotoxicity was absent in cyclising (Group A) and in Group C compounds. Toxicity, expressed by comparison with 4-hydroxyanisole (4HA) (IC50 = 11.7 microM), ranged between 0.36 (4-hydroxybenzyl alcohol) and 1.07 (3-(4-hydroxyphenyl)propanol) for Group B compounds, and be-tween 0.83 (4-ethoxyphenol) and 2.08 (4-(2-hydroxyethylthio)phenol) for groups D, E and F. Addition of glutathione to the toxicity assay system abrogated the cytotoxic action and, on the basis of spectrophotometric data, this is ascribed to the prevention of cellular thiol depletion by the ortho-quinone products of tyrosinase oxidation of the phenolic substrates. The lack of toxicity of the group C compounds may be due to the inability of their derived quinones to gain access to the cells. Addition of catalase or deferoxamine to the incubation medium was without effect on tyrosinase-dependent toxicity.

Selective tumor uptake of a boronated porphyrin in an animal model of cerebral glioma.

The prognosis for patients with high-grade cerebral glioma is poor. Most treatment failures are due to local recurrence of tumor, indicating that a more aggressive local therapy could be beneficial. Adjuvant treatments such as porphyrin-sensitized photodynamic therapy (PDT) or boron neutron capture therapy (BNCT) have the potential to control local recurrence. The selective tumor uptake of a boronated porphyrin was studied in CBA mice bearing an implanted intracerebral glioma. Biopsy samples of tumor, normal brain, and blood were analyzed by a fluorometric assay following intraperitoneal and intravenous administration of boronated protoporphyrin (BOPP). This compound was selectively localized to tumor at ratios as high as 400:1 relative to normal brain. Confocal laser scanning microscopy of glioma cells in vitro and in vivo showed that BOPP was localized within mitochondria and excluded from the nucleus of these cells. This discrete subcellular localization was confirmed by density gradient ultracentrifugation after homogenization of mouse tumor biopsies. The selective discrete localization of these compounds within the tumor suggests that this compound may be used as a dual PDT/BNCT sensitizer.

In vitro assessment of the structure-activity relationship of tyrosinase-dependent cytotoxicity of a series of substituted phenols.

The rate of oxidation by purified mushroom tyrosinase of 30 compounds was measured by oximetry, and the tyrosinase-dependent cytotoxicity of each estimated in an in vitro assay using exposure of non-melanogenic cells to the agents in the presence and absence of tyrosinase. Cytotoxicity was estimated by immediate inhibition of DNA synthesis; 4-hydroxyanisole was used as the reference material. Compounds that were not oxidized by tyrosinase were found to be non-toxic but there was no direct relationship between the rate of oxidation and the relative cytotoxicity of those materials that acted as substrates for the enzyme. Thioethers were found to be more cytotoxic than the corresponding phenoxyethers. This was partly due to their greater rate of oxidation by tyrosinase and, in the case of propylthiophenol, the consequence of higher effective toxicity of the lipophilic species. The optimum chain length for the side chain of the oxyethers was three saturated carbon atoms and the toxicity appeared to be influenced by the lipophilicity of the compounds, possibly reflecting the relative lipid solubility of the putative toxic ortho-quinones generated from them. The maximum tyrosinase-dependent toxicity observed was in the range 5-6 times the relative toxicity of 4-hydroxyanisole. Sulphinyl and sulphonyl derivatives were inactive. In addition to oxyethers and thioethers, esters and glycosides of oxyethers were also examined and were found to be toxic in the presence of tyrosinase when hydrolysed. The succinates were found to be oxidized and toxic in our test system, suggesting that they rapidly underwent spontaneous hydrolysis. Oximetry data suggest that slight spontaneous hydrolysis of the other compounds occurs but they were not toxic in our assay. Ring-methylated phenoxyethers were oxidized relatively slowly and were non-toxic. Fluorine-substituted phenoxyethers were oxidized slightly more rapidly and exhibited clear toxicity in our system. Sesamol was oxidized to a black pigment but was non-toxic in our assay. A water-soluble vitamin E derivative was not oxidized and was non-toxic. Allyl hydroquinone was not oxidized but exhibited significant direct toxicity.

Effects of perindopril on tissue angiotensin-converting enzyme activity demonstrated by quantitative in vitro autoradiography.

Inhibition of plasma angiotensin II generation does not fully explain the chronic hypotensive effects of angiotensin-converting enzyme (ACE) inhibitors. Therefore, the pattern of tissue ACE inhibition in rats was studied after oral administration of perindopril, a new ACE inhibitor. Tissue ACE was measured by quantitative in vitro autoradiography using [125I]-351A as a radioligand and compared with plasma ACE and the pressor response to intravenous (i.v.) angiotensin I. Following oral perindopril (1 mg/kg), plasma ACE activity was acutely reduced, but recovered over 24 h. The peak concentration of plasma perindoprilic acid, the active diacid of perindopril, occurred at 1 h, and the drug was undetectable by 24 h. The pressor response to i.v. angiotensin I was inhibited by 95% at 4 h and had not fully recovered by 24 h. Four hours after oral administration of perindopril, ACE was markedly inhibited in the proximal tubules of the kidney (24% control), lung parenchyma (10%), and aortic wall (18%) (p less than 0.01). At 24 h, ACE in these tissues had only partially recovered (32-63%). ACE was also identified in vascular endothelium of organs, including the lung, kidney, and testis; in these sites, vascular ACE showed a pattern of inhibition similar to that of aortic ACE. In contrast, ACE in testicular seminiferous tubules was unaffected by perindopril. These results demonstrate a prolonged effect of ACE inhibitors on tissue ACE that may better explain the time course of these drugs than the changes in plasma ACE or plasma levels of the drug.

Dialysis osteomalacia: a possible role for zirconium as well as aluminium.

Six patients with dialysis osteomalacia were studied before and after treatment with desferrioxamine. Before treatment, all six had severe osteomalacia with histochemical evidence of metals at the mineralization front, confirmed by energy dispersive X-ray microanalysis to include Al, Zr and Fe. Zr was not detected by histological staining in patients without dialysis osteomalacia. After treatment a decrease of Al and Zr was associated with improvement in clinical, biochemical, radiological and histological parameters. These observations suggest the possibility of a role for metals other than Al in the pathogenesis of dialysis osteomalacia.

Dietetics training for American Indians and Alaskan natives.

The response to training has been enthusiastic. Even in these times of limited funds, applications to attend training exceed the available space. From the first class in October 1968 through September 1982, nearly 1,300 Indian and Alaskan native hospital food service employees and employees representing tribal programs throughout the country have received training from the courses and workshops provided by the Nutrition and Dietetics Training Program. With the increasing involvement of Native Americans in their own health care programs, the need for training in foods and nutrition will continue.

Intensity of guitar playing as a function of auditory feedback.

Subjects played an electric guitar while auditory feedback was attenuated or amplified at seven sidetone levels varying 10-dB steps around a comfortable listening level. The sidetone signal was presented in quiet (experiment I) and several levels of white noise (experiment II). Subjects compensated for feedback changes, demonstrating a sidetone amplification as well as a Lombard effect. The similarity of these results to those found previously for speech suggests that guitar playing can be a useful analog for the function of auditory feedback in speech production. Unlike previous findings for speech, the sidetone-amplification effect was not potentiated by masking, consistent with a hypothesis that potentiation in speech is attributable to interference with bone conduction caused by the masking noise.