Age adjusted Charlson Co-morbidity Index is an independent predictor of mortality over long-term follow-up in infective endocarditis.

BACKGROUND: Infective endocarditis (IE) is associated with high morbidity and mortality. The epidemiology of IE is changing, affecting more elderly patients with increased medical comorbidities. We aimed to assess the ability of the age adjusted Charlson Co-morbidity Index (ACCI) to predict early and late outcomes. METHODS: Between 1998 and 2010, adult patients with definite IE according to the modified Duke criteria were identified. The primary outcome was in-hospital and all-cause mortality. The secondary outcome was predictors of the primary outcome incorporating ACCI. RESULTS: 148 patients with IE were followed up for a mean of 3.8 ± 3 years. The mean age was 57 ± 17 years and 66% were male. In-hospital mortality and all-cause mortality were 24 and 47% respectively. Comorbid conditions included diabetes mellitus (DM) (21%); ischaemic heart disease (16%); heart failure (HF) (14%); renal failure (eGFR <60 ml/min/1.73 m(2)) (19%); and anaemia (64%). The most common causative organism was Staphylococcus aureus (53%). ACCI was >3 in 59% of patients. Cardiac surgery was performed in 45% of patients. On Cox regression analysis, ACCI >3 (HR=3.0 [1.5-6.0], p<0.002), new onset HF (HR=2.2 [1.3-3.6], p<0.003), anaemia (HR=1.8 [1.1-3.2], p=0.04) and age-per decade (HR=1.4 [1.1-1.7]. p=0.004) were independently associated with all-cause mortality. ACCI >3 was the strongest predictor of in-hospital mortality (OR=8.4 [2.8-24], p<0.001). Of the individual ACCI components, prior HF, DM with complications and metastatic disease were independent predictors of all-cause mortality. CONCLUSION: In-hospital and all-cause mortality of IE remain high. An ACCI >3 was a strong predictor of mortality, in addition to age, new HF and anaemia.

Progression of aortic stenosis in elderly patients over long-term follow up.

BACKGROUND: The natural history of aortic stenosis (AS) in elderly patients remains poorly defined. In an elderly cohort over long-term follow-up, we assessed: 1) rates and predictors of hemodynamic progression and 2) composite aortic valve replacement (AVR) or death endpoint. METHODS: Consecutive Department of Veterans' Affairs patients with AS (>60 years) were prospectively enrolled between 1988 and 1994 (n=239) and followed until 2008. Patients with ≥ 2 trans-thoracic echocardiograms >6 months apart were included in the progression analysis (n=147). Baseline demographics, comorbidities and echocardiography parameters were recorded. Follow-up was censored at AVR/death. RESULTS: The age of patients was 73 ± 6 years; 82% were male. Baseline AS severity was mild (67%), moderate (23%) and severe (10%). Follow-up was 6.5 ± 4 years (range: 1-17 years). Annualized mean aortic valve gradient progression rates were: mild AS 4 ± 4 mmHg/year; moderate AS 6 ± 5 mmHg/year and severe AS 10 ± 8 mmHg/year (p<0.001). Five-year event-free survival was 66 ± 5%, 23 ± 7% and 20 ± 10% for mild, moderate and severe AS respectively. Progression to severe AS occurred in 35% and 74% of patients with mild and moderate AS respectively. Independent predictors of rapid progression were: baseline AS severity (per grade) (OR 2.6, p=0.001), aortic valve calcification (per grade) (OR 2.1, p=0.01), severe renal impairment (OR 4.0, p=0.04) and anemia (OR 2.3, p=0.05). CONCLUSIONS: In elderly patients, hemodynamic progression of AS is predicted by AS severity, renal function, aortic valve calcification and history of anemia. These factors identify patients at high risk of rapid hemodynamic progression, for whom more frequent clinical and echocardiographic surveillance is advisable.

Global longitudinal strain is a strong independent predictor of all-cause mortality in patients with aortic stenosis.

AIMS: To assess the capacity of global longitudinal strain (GLS) in patients with aortic stenosis (AS) to (i) detect the subclinical left ventricular (LV) dysfunction [LV ejection fraction (LVEF) ≥50% patients]; (ii) predict all-cause mortality and major adverse cardiac events (MACE) (all patients), and (iii) provide incremental prognostic information over current risk markers. METHODS AND RESULTS: Patients with AS (n = 146) and age-matched controls (n = 12) underwent baseline echocardiography to assess AS severity, conventional LV parameters and GLS via speckle tracking echocardiography. Baseline demographics, symptom severity class and comorbidities were recorded. Outcomes were identified via hospital record review and subject/physician interview. The mean age was 75 ± 11, 62% were male. The baseline aortic valve (AV) area was 1.0 ± 0.4 cm(2) and LVEF was 59 ± 11%. In patients with a normal LVEF (n = 122), the baseline GLS was controls -21 ± 2%, mild AS -18 ± 3%, moderate AS -17 ± 3% and severe AS -15 ± 3% (P< 0.001). GLS correlated with the LV mass index, LVEF, AS severity, and symptom class (P< 0.05). During a median follow-up of 2.1 (inter-quartile range: 1.8-2.4) years, there were 20 deaths and 101 MACE. Unadjusted hazard ratios (HRs) for GLS (per %) were all-cause mortality (HR: 1.42, P< 0.001) and MACE (HR: 1.09, P< 0.001). After adjustment for clinical and echocardiographic variables, GLS remained a strong independent predictor of all-cause mortality (HR: 1.38, P< 0.001). CONCLUSIONS: GLS detects subclinical dysfunction and has incremental prognostic value over traditional risk markers including haemodynamic severity, symptom class, and LVEF in patients with AS. Incorporation of GLS into risk models may improve the identification of the optimal timing for AV replacement.

Effect of drug-polymer binary mixtures on the in-vitro release of ibuprofen from transdermal drug-in-adhesive layers.

We report on the formation of eutectic mixtures of ibuprofen using two different polymers together with investigations on the in-vitro release of ibuprofen from drug-in-adhesive layers. Ibuprofen, literature melting point (m.p.) = 73.5-76.5°C, was tested together with Pluronic F127, literature m.p. = 54.4-60.5°C, and polyethylene glycol 1000 (PEG 1000), literature m.p. = 37-40.9°C, as second components in binary mixtures, incorporated into an acrylic adhesive, either as solid physical mixtures (PM) or molten mixtures (MM). Studies of how the type of mixture preparation (PM versus MM) and the ratio of components in binary mixtures affecting the in-vitro drug release of ibuprofen, compared with ibuprofen-adhesive layers without polymer addition were conducted. Ibuprofen release did not improve using the eutectic composition with Pluronic F127, possibly due to increased ibuprofen solubilisation in the adhesive and a subsequent decrease in the thermodynamic activity of the formulation. A significant increase in ibuprofen release (P < 0.05) was shown for compositions adjacent to the eutectic one, with ibuprofen: Pluronic F127 (40:60) and ibuprofen: PEG 1000 (20:80, 25:75, 30:70), from both PM- and MM-adhesive formulations, compared to the ibuprofen-adhesive formulations.

Percutaneous balloon valvuloplasty of a stenosed mitral bioprosthesis.

A 62-year-old woman with disabling mitral prosthetic stenosis underwent percutaneous balloon valvuloplasty. The transvalvular gradient preoperatively was 22 mm Hg and was reduced to 6 mm Hg after the valvuloplasty. the valve area was increased from an initial value of 0.77 cm2 to 1.53 cm2. No complications occurred related to the Further studies are necessary to ascertain the indications and long-term results of percutaneous valvuloplasty on bioprosthesis in the mitral position.

Changes in muntjac fibroblasts associated with the acquisition of cadmium resistance. A pre-resistance, transitional and post-resistance study.

A series of cell lines with different levels of resistance to continuous cadmium exposure has been developed from an immortal but non-transformed muntjac fibroblast cell line. Concentrations accepted in their culture medium range from 0.1 microM for the cadmium sensitive parent line to 5 microM for the intermediate "cadmium-tolerant" line, to 5, 10, 20 and 50 microM for the four "cadmium-resistant" lines. The present paper follows the morphological changes which accompanied the development of resistance through a 20-month pre-resistance period, a relatively abrupt 6-week transitional period and a 3-year post-resistance period, during which time levels of cadmium resistance were increased. Initial changes which led to the cadmium-tolerant CR5 cell line included (i) increased efficiency in autophagocytosing damaged cell components and in ridding the cell of residual waste materials, (ii) a reduction in fluid filled vacuoles and (iii) improved recycling and/or replacement of cadmium-damaged cell membrane. With the advent of cadmium resistance the intracellular damage necessitating these activities disappeared, yet the series of changes which occurred included a massive build-up of Golgi and the appearance of a trans-Golgi tubular network in addition to cytoskeletal and membrane changes. Though metallothionein levels are greater in the cadmium-resistant variants, their increase appears inadequate on their own to account for the high levels of resistance. The post-resistance changes which accompanied each step-up in cadmium resistance included further membrane and glycocalyx changes, in addition to continued increases in Golgi bodies and tubular network. This paper details the morphological changes which occurred throughout the 5-year period, tests the direct dependence of each on the presence of cadmium and examines their possible contribution to a cadmium protective mechanism.

Cadmium-induced multistep transformation of cultured Indian muntjac skin fibroblasts.

During the past five years we have made a series of cadmium-transformed and resistant fibroblast cell lines by continuous low-level exposure to cadmium. In the present paper we describe the use of four of these lines with varying degrees of transformation to investigate the multistep nature of cadmium carcinogenesis. These include: (a) M cell, an immortal but nontransformed muntjac skin fibroblast line; (b) CCR5, a morphologically transformed and cadmium-resistant line derived from M cells after 20-months continuous exposure to small step-wise increases in cadmium; (c) SCR5, a tumorigenic line derived by selection (in the absence of cadmium) of rapidly growing CCR5 agar colonies; (d) T1, a line derived from an SCR5 tumour growing in a nude mouse. We have compared the morphological characteristics of the four cell lines using light and electron microscopy and evaluated their ability to grow in liquid culture, soft agar and nude mice. We have also examined the changes which have occurred in their cytoskeletons and extracellular matrices using fluorescent antibodies to actin, tubulin and fibronectin and related these to the strength of their cell-cell and cell-substrate attachments and to their levels of transformation and tumorigenesis. We have shown that, while some changes occur in a single step (e.g. intracellular cytoskeletal changes), others are gradual (e.g. changes in extracellular matrix, focus formation and ability to grow in soft agar). We conclude that continuous exposure to low levels of cadmium can initiate growth and structural changes which subsequently lead to cell transformation and tumorigenesis on the removal of cadmium. Though change with cadmium was slow, many of the transformed characteristics are similar to those reported for viral and chemically transformed cells.

Mechanism of cadmium resistance in cultured Indian muntjac fibroblasts.

The mechanism by which Indian muntjac fibroblasts (M cells) became resistant to the toxic effects of cadmium was investigated. The resistant lines, CCR5 and CCR20, were approximately 8-fold and 18-fold more resistant to acute cadmium exposures than the parental M cell line. Examination of the roles of metallothionein, glutathione and membrane permeability indicates that although resistance may be multifactorial, the major change involves decreased uptake of cadmium by resistant cells.

A growth factor for hepatocytes is inactivated by sub-lethal gamma-irradiation in vivo or in vitro.

Serum from partially hepatectomized rats promoted DNA synthesis in primary adult rat hepatocyte cultures. If the rats had been exposed to sub-lethal gamma-irradiation immediately following operation or if their serum, collected at 3 h, was exposed to irradiation in vitro, the growth-promoting activity was destroyed. Prostaglandin E2 also stimulated DNA synthesis in the cultures; if PGE2 was irradiated in serum from intact or partially hepatectomized rats its growth-promoting activity was markedly diminished.

The mutagenic and carcinogenic properties of three second generation antitumour platinum compounds: a comparison with cisplatin.

The cytotoxicity, mutagenicity and transforming potentials of three second generation platinum compounds have been investigated in mammalian cells. All the compounds showed positive response in two assay systems in Chinese hamster V 79 cells, i.e. measurement of mutation induction at the HGPRT locus and of DNA damage as indicated by sister chromatid exchange frequencies. At equitoxic doses, the compounds in order of decreasing mutagenicities were cisplatin, spiroplatin, carboplatin and iproplatin. The BHK transformation assay reflected a similar order in the potential carcinogenicity of the drugs. Cisplatin was highly carcinogenic, followed by spiroplatin. In comparison, carboplatin and iproplatin were potentially weak carcinogens.

Serum from partially hepatectomized rats induces primary hepatocytes to enter S phase: a role for prostaglandins?

Serum obtained from partially hepatectomized rats 1, 3 or 24 h after operation was more effective in stimulating DNA synthesis in primary adult rat hepatocytes than serum from sham-operated rats; exposure to the serum for 2 h was sufficient to promote growth. Serum from the partially hepatectomized rats contained elevated levels of PGE2 and PGF2 alpha; it promoted hepatocytes to release prostaglandins into their culture medium. Growth-promoting effects of the serum and its capacity to elicit prostaglandin release into the culture medium were inhibited by 0.1 mM-indomethacin or 1 mM-aspirin. 0.1 mM-indomethacin also prevented DNA synthesis if the inhibitor were added 4 h after growth had been initiated by serum from partially hepatectomized rats, suggesting that prostaglandins continue to be important for the maintenance of hepatocyte growth for at least 6 h.

Parallel development of cadmium resistance and in vitro transformation in cultured Indian muntjac cells.

The development of cadmium resistance in an Indian muntjac cell line has been investigated. The parent cell line is highly sensitive to cadmium ions. Resistance was obtained by continuous growth of cells in low levels of cadmium with stepwise increments. Four cell lines were developed with resistances of between 50- and 200-fold greater than that of the parental line. Early in the development of resistance an unstable cell line displaying extensive chromosomal rearrangement and an elevated sister chromatid exchange frequency was identified. The more stable resistant lines produced from this original cell line have normal karyotypes. Having passed through the initial period of genome rearrangement the resultant cells acquired several characteristics of morphologically transformed cells. It is concluded that long-term exposure to low levels of cadmium can transform cells in vitro concurrently with their acquiring cadmium resistance.

Regulation of the proliferation of primary rat hepatocytes by eicosanoids.

DNA synthesis in primary adult rat hepatocyte cultures was promoted by epidermal growth factor (EGF), arachidonic acid, and prostaglandins E2 and F2 alpha (PGE2 and PGF2 alpha). Growth promotion by EGF was blocked by 0.1 mM indomethacin and 1 mM aspirin, without affecting cell viability. If verapamil was present in the medium when EGF was added, the growth response was inhibited. Hepatocytes stimulated by EGF or arachidonic acid released PGE2 and PGF2 alpha into the culture medium. This was diminished if 0.1 mM indomethacin was also in the medium. The importance of autocrine regulation of hepatocyte growth by prostaglandins is discussed.

Cadmium induced changes in cell organelles: an ultrastructural study using cadmium sensitive and resistant muntjac fibroblast cell lines.

A detailed electron microscopy study of cadmium sensitive and resistant muntjac fibroblast cell lines has identified a wide range of intracellular damage following exposure to cadmium. Damaged organelles included cell membrane, mitochondria, Golgi cisternae and tubular network, chromatin, nucleoli, microfilaments and ribosomes. Although cell membrane damage was generally the earliest indication of adverse cadmium action, particularly with continuous cadmium exposures, cells could tolerate extensive membrane loss. Mitochondrial distortion and some damage to Golgi was also tolerated. The turning point at which cadmium became lethal was generally marked by a cascade of events which included damage to both nuclear and cytoplasmic components. These results for fibroblasts are discussed and compared with damage reported in other types of cells.

The radiosensitivity of rat thymocytes.

gamma-Irradiation in vitro apparently blocked a plasma-membrane associated, superoxide-producing, NADPH oxidase in rat thymocytes. Differential centrifugation of the mixed thymocytes indicated the smaller lymphocytes (approx. 6 microns diameter) to be the radiosensitive population. The oxidase system co-isolated in part with thymus nuclei and could be solubilized by detergent treatment [Bellavite, Jones, Cross, Papini & Rossi (1984) Biochem. J. 223, 639-648]. Endogenous NADPH was the rate-limiting component for superoxide formation in vitro. The level of NADPH was lowered by gamma-irradiation, an effect mimicked by GSSG in the presence of 50 microM-ZnCl2 to inhibit GSSG reductase. These findings are suggested as the metabolic basis for interphase death of small lymphocytes exposed to ionizing radiation.

Protein kinase activities in rat liver nuclei: effects of age and partial hepatectomy.

Selective substrates and inhibitors have been used to measure kinases phosphorylating endogenous proteins in rat liver nuclei during growth and regeneration after partial hepatectomy. Peaks in activity were found at 5, 22, and 29 hours after partial hepatectomy. Administration of alpha 1 and beta adrenergic blockers suggested that the Be2+ sensitive and cyclic AMP-dependent protein kinases were interdependently regulated by Ca2+ and cyclic AMP.

Novobiocin inhibition of DNA excision repair may occur through effects on mitochondrial structure and ATP metabolism, not on repair topoisomerases.

Novobiocin inhibits DNA topoisomerases. It also inhibits excision repair of DNA photodamage, blocking both repair synthesis and the earlier step of incision at u.v. damage sites (as measured by the accumulation of DNA strand breaks in u.v.-irradiated interphase cells treated with DNA synthesis inhibitors such as hydroxyurea or cytosine arabinoside). It has been supposed, therefore, that novobiocin affects repair by blocking a putative topoisomerase step prior to incision. But we find that novobiocin also has a marked dose- and time-dependent effect on mitochondria: in cells exposed to novobiocin, mitochondria swell and their cristae become disrupted, and the intracellular ATP:ADP ratio is lowered, though the membrane potential is maintained as judged by rhodamine 123 fluorescence. Mitotic cells are more resistant to mitochondrial disruption by novobiocin than are interphase cells. This correlates with a relative resistance of u.v.-irradiated mitotic cells to the inhibition of incision by novobiocin. The chromosomal decondensation that results from the accumulation of DNA breaks due to incision when u.v.-irradiated mitotic cells are treated with hydroxyurea and cytosine arabinoside is largely suppressed by novobiocin. Furthermore, the suppression of induced strand break accumulation is partly due to a suppression by novobiocin of the uptake and phosphorylation of cytosine arabinoside; breaks accumulated in u.v.-irradiated cells in the presence of aphidicolin, an inhibitor of DNA polymerase alpha that does not require phosphorylation, are less novobiocin-sensitive. We conclude that the effects of novobiocin on excision repair are more likely to be due to a non-specific effect on ATP metabolism than to a specific effect on a repair-related topoisomerase.

The ability of bifunctional and monofunctional pyrrole compounds to induce sister-chromatid exchange (SCE) in human lymphocytes and mutations in Salmonella typhimurium.

The ability to induce sister-chromatid exchange (SCE) in human lymphocytes and mutations in Salmonella typhimurium has been assessed for 4 pyrrole compounds. Three of the compounds, 2,3-bishydroxymethyl-1-methylpyrrole (BHMP), 2-hydroxymethyl-1-methylpyrrole (2HMP) and 3-hydroxymethyl-1-methylpyrrole (3HMP) are synthetic pyrrolic alcohols; the fourth compound, dehydroretronecine (DHR) is a metabolite of several naturally occurring pyrrolizidine alkaloids. The activity of these compounds was compared with that of mitomycin C (MMC) and decarbamoyl mitomycin C (DCMMC), chemicals related structurally to the pyrrole compounds. All 6 compounds caused an increase in the numbers of SCEs. Whereas the bifunctional pyrroles, DHR and BHMP, and the mitomycins, MMC and DCMMC, increased levels of SCEs by 8-12 times control levels, the monofunctional pyrroles gave increases of only 2 times. Three of the 4 pyrrole compounds (DHR, BHMP and 3HMP) induced mutations in the Salmonella typhimurium base substitution strain TA92, the fourth (2HMP) was not found to be mutagenic in any of the 8 strains used. The mitomycins induced mutations in the frameshift strain TA94 in addition to the base substitution strain TA92, with DCMMC always more mutagenic and less cytotoxic than MMC. All bifunctional compounds induced more mutations and were less cytotoxic in strains containing an efficient excision-repair system. With the pyrrole compounds numbers of SCEs and mutations were only increased when using chemical concentrations significantly higher than those required for the mitomycins: more than twice as high to produce significant numbers of SCEs and more than 100 times as high to produce equal numbers of mutations.