Effect of thalidomide on the skeletal muscle in experimental heart failure

BACKGROUND: Tumour Necrosis Factor alpha (TNFalpha) has been shown to contribute to heart failure (CHF) progression. AIMS: We have tried to antagonise the detrimental effects of TNFalpha on skeletal muscle apoptosis, by using thalidomide, a drug that inhibits its biosynthesis. METHODS: CHF was induced in 20 rats by injecting monocrotaline, which determines right ventricle (RV) failure. After 2 weeks, when CHF developed, 12 rats were treated with thalidomide 3.5.mg/kg per day for 2 weeks. Eight had saline and served as CHF controls. RESULTS: Thalidomide failed to decrease TNFalpha and its second messenger sphingosine (SPH), but was able to prevent the shift toward the fast myosin heavy chains. In the Tibialis Anterior muscle of the thalidomide group, the degree of atrophy, the number of apoptotic nuclei and the levels of caspases, were similar to those of the CHF controls. CONCLUSIONS: Thalidomide, at the doses used in this study, which are the same employed for the treatment of tubercolosis, leprosy, AIDS and cancer in humans, did not lower either TNFalpha or SPH and only marginally influenced the apoptosis-induced muscle atrophy. Since other TNFalpha blockers are under investigation for improving the clinical status of patients with CHF, the present data could be relevant in the design of randomised clinical trials in humans.     

Cell cycle deregulation in liver lesions of rats with and without genetic predisposition to hepatocarcinogenesis

Preneoplastic and neoplastic hepatocytes undergo c-Myc up-regulation and overgrowth in rats genetically susceptible to hepatocarcinogenesis, but not in resistant rats. Because c-Myc regulates the pRb-E2F pathway, we evaluated cell cycle gene expression in neoplastic nodules and hepatocellular carcinomas (HCCs), induced by initiation/selection (IS) protocols 40 and 70 weeks after diethylnitrosamine treatment, in susceptible Fisher 344 (F344) rats, and resistant Wistar and Brown Norway (BN) rats. No interstrain differences in gene expression occurred in normal liver. Overexpression of c-myc, Cyclins D1, E, and A, and E2F1 genes, at messenger RNA (mRNA) and protein levels, rise in Cyclin D1-CDK4, Cyclin E-CDK2, and E2F1-DP1 complexes, and pRb hyperphosphorylation occurred in nodules and HCCs of F344 rats. Expression of Cdk4, Cdk2, p16(INK4A), and p27(KIP1) did not change. In nodules and/or HCCs of Wistar and BN rats, low or no increases in c-myc, Cyclins D1, E, and A, and E2F1 expression, and Cyclin-CDKs complex formation were associated with p16(INK4A) overexpression and pRb hypophosphorylation. In conclusion, these results suggest deregulation of G1 and S phases in liver lesions of susceptible rats and block of G1-S transition in lesions of resistant strains, which explains their low progression capacity.     

Motion sickness and postoperative vomiting in children

BACKGROUND: Motion sickness is considered an important risk factor for postoperative nausea and vomiting in children. The aim of this study was to verify the impact of motion sickness on the incidence of vomiting after routine surgery in children, and to compare the incidence of vomiting, after combined regional/general anaesthesia, using either halothane or sevoflurane. METHODS: We prospectively studied 420 children (369 males and 51 females) who received general anaesthesia and inguinal field block for common paediatric surgery. The children were randomly allocated into one of two groups (halothane or sevoflurane). In the 200 children in the first group (H), general anaesthesia was induced and maintained with halothane, whereas in the 220 children in the second group (S), anaesthesia was induced and maintained with sevoflurane. RESULTS: There were 79 children with a prior history of motion sickness (MS+) and 341 without such a history (MS-). In the MS+ population, the incidence of vomiting was similar in both H and S groups, being around 33%. However, repeated episodes of vomiting in MS+ children were more frequent when halothane was used. In the MS- group, the incidence of vomiting was significantly greater in the H group (19%) than in the S group (8%). CONCLUSIONS: In the postoperative period, we found that MS+ children vomit more than MS- children, regardless of the inhalation anaesthetic used. However, MS- children displayed a higher incidence of vomiting when halothane was used rather than sevoflurane.     

Apoptosis in the skeletal muscle of patients with heart failure: investigation of clinical and biochemical changes

OBJECTIVE—To investigate the contribution of apoptosis in the development of the skeletal myopathy in chronic heart failure.
DESIGN—The electrophoretic pattern of myosin heavy chains (MHC), fibre cross sectional area, number of in situ nick end labelling (TUNEL) positive apoptotic myocyte nuclei, and the tissue levels of caspase-3, Bcl-2, and ubiquitin were determined in biopsies taken from the vastus lateralis muscle. The study involved nine patients with severe chronic heart failure caused by ischaemic heart disease and hibernating myocardium and five controls.
RESULTS—In chronic heart failure patients the vastus lateralis showed a significant increase of MHC_2a and MHC_2b and a greater degree of fibre atrophy, as demonstrated by the decreased cross sectional area. There was also an increased number of TUNEL positive apoptotic myocyte nuclei. Tissue concentrations of Bcl-2 were decreased, while those of caspase-3 and ubiquitin were increased. Peak oxygen consumption (VO₂) was negatively correlated with the number of TUNEL positive nuclei and the fibre cross sectional area. There was a correlation between the number of apoptotic nuclei and the fibre cross sectional area, but no correlation between myosin heavy chains and number of apoptotic nuclei.
CONCLUSIONS—Myocyte apoptosis occurs in the skeletal muscle of patients with chronic heart failure, and its magnitude is associated with the severity of exercise capacity limitation and the degree of muscle atrophy. Muscle atrophy contributes to the limitation of exercise capacity, together with the increased synthesis of fast, more fatiguable myosin heavy chains.


Keywords: apoptosis; chronic heart failure; exercise capacity; myosin heavy chains     

A phase I-II study on a gemcitabine-cyclophosphamide-fluorouracil/folinic acid triplet combination in anthracycline- and taxane-refractory breast cancer patients.

PURPOSE: To define the cyclophosphamide (CTX) maximal tolerated dose when combined with fixed doses of gemcitabine, fluorouracil (5-FU) and folinic acid (leucovorin, LFA) in metastatic breast cancer patients pretreated with anthracyclines and taxanes. METHODS: Metastatic breast cancer patients aged < or = 75 years, with ECOG performance status 0-2, were eligible, provided that they had received previous anthracycline- and taxane-based chemotherapy for the advanced disease. Chemotherapy consisted of gemcitabine 1,000 mg/m(2), 5-FU 425 mg/m(2), LFA 100 mg/m(2) and escalating doses of CTX, starting from 500 mg/m(2), on days 1 and 8 every 3 weeks. The dose escalation was stopped if dose-limiting toxicity (DLT) occurred in > 33% of patients of a given cohort. After the definition of DLT, a further escalation with the addition of granulocyte colony-stimulating factor (G-CSF; on days 3-5 and 10-12) was planned. RESULTS: Since March 1999, 69 patients have entered this trial through seven different cohorts. The dose escalation was stopped at the CTX dose of 600 mg/m(2) since 3/6 patients showed DLT. A further dose escalation was attempted in the presence of G-CSF support. A CTX dose of 800 mg/m(2) proved to be safe and was chosen for the phase II. A total of 33 patients were treated at this dose level. The treatment was fairly well tolerated, grade 3-4 neutropenia and thrombocytopenia occurring in 38 and 16% of patients, respectively. No cases of sepsis or bleeding were registered. Four patients required a packed red blood cell transfusion. Severe nonhematologic toxicity was also uncommon, occurring in 10 patients. Three complete and 24 partial responses were recorded for an overall response rate of 38% (95% CI = 26-50). Two complete and 12 partial responses were recorded in the 33 patients treated in the phase II for an overall response rate (ORR) of 42% (95% CI = 25-61). CONCLUSIONS: The gemcitabine-CTX-5-FU/LFA combination is a well-tolerated treatment for poor-prognosis breast cancer patients with previous exposure to anthracyclines and taxanes. With the addition of G-CSF, a cumulative CTX dose of 1,600 mg/m(2) can be safely delivered every 3 weeks. The evidence of an ORR approaching 40% is very promising and justifies further evaluations in this subset of patients.     

Low-dose oral methotrexate and cyclophosphamide in metastatic breast cancer: antitumor activity and correlation with vascular endothelial growth factor levels.

BACKGROUND: Anticancer chemotherapy is thought to be effective by means of direct cytotoxicity on tumor cells. Alternative mechanisms of efficacy have been ascribed to several common anticancer agents, including cyclophosphamide (CTX), methotrexate (MTX), anthracyclines and taxanes, postulating an antiangiogenic activity. PATIENTS AND METHODS: We evaluated the clinical efficacy and impact on serum vascular endothelial growth factor (VEGF) levels of low-dose oral MTX and CTX in patients with metastatic breast cancer. MTX was administered 2.5 mg bd on days 1 and 2 each week and CTX 50 mg/day administered continuously. RESULTS: Sixty-four patients were enrolled, 63 were evaluable: Eastern Cooperative Oncology Group (ECOG) performance status 0-1, > or =2 sites of metastatic disease (n = 50 patients), progressive disease at study entry (n = 51), 1 regimen for metastatic disease (n = 32) and > or =2 regimens (n = 20). Among the 63 evaluable patients, there were two complete remissions (CR), 10 partial remissions (PR) for an overall response rate of 19.0% (95% CI 10.2% to 30.9%) and an overall clinical benefit (CR+ PR+ stable disease >24 weeks) of 31.7% (95% CI 20.6% to 44.7%). Grade > or =2 leucopenia was registered in only 13 patients. The median serum VEGF level for the subgroup of patients on treatment for at least 2 months decreased with treatment from 315 pg/ml (95% CI 245 to 435) at baseline to 248 pg/ml (95% CI 205 to 311) at 2 months (P <0.001). Both responders and non-responders showed similar reductions in serum VEGF (P = 0.78). After 6 months patients still on treatment had a median VEGF level of 195 pg/ml (95% CI 96 to 355), which was significantly lower than the median baseline values (P = 0.001). CONCLUSIONS: Continuously low-dose CTX and MTX is minimally toxic and effective in heavily pretreated breast cancer patients. A drop in VEGF was associated with the treatment and so alternative hypotheses, other than that of direct toxicity on tumor cells, must be favored when trying to explain the anticancer effect.     

Myocardial injury revealed by plasma troponin I in breast cancer treated with high-dose chemotherapy.

BACKGROUND: High-dose chemotherapy (HDC) has been widely utilized in high-risk breast cancer, but it may induce cardiac toxicity. Cardiac dysfunction may become evident weeks or months after HDC and, to date, no early markers of myocardial injury that are able to predict late ventricular impairment are available. We investigated the role of plasma troponin I (TnI) in this setting. PATIENTS AND METHODS: We measured TnI plasma concentration after HDC in 211 high-risk breast cancer women (46 +/- 11 years, mean +/- SD). According to TnI value (< 0.5 or > or = 0.5 ng/ml), patients were allocated into a troponin positive (TnI+; n = 70) and a troponin negative (TnI-; n = 141) group. All patients underwent left ventricular ejection fraction (LVEF, Echo) examination during the following 12 months. RESULTS: LVEF progressively decreased in the TnI+ group but not in the TnI- group. In TnI+ patients a close relationship between the TnI increase, as well as the number of positive TnI assays, and the maximal LVEF decrement, was found (r = -0.92, P < 0.0001 and r = -0.93, P < 0.0001, respectively). CONCLUSIONS: In our population, the elevation of TnI soon after HDC accurately predicts the development of future LVEF depression. In this setting, TnI can be considered a sensitive and reliable marker of myocardial damage with relevant clinical and prognostic implications.     

Different Effects of Regenerative and Direct Mitogenic Stimuli on the Growth of Initiated Cells in the Resistant Hepatocyte Model

The possible mechanism(s) responsible for the different effects exerted by proliferative stimuli of different nature on the appearance of enzyme-altered hepatic foci, were investigated in male Wistar rats. Rats given an initiating dose of diethylnitrosamine (150 mg/kg body weight) were fed a diet containing 0.03% acetylaminofluorene for 2 weeks. Between the first and the second week, cell proliferation was induced by a proliferative stimulus of compensatory type (partial hepatectomy) or by a direct mitogenic stimulus (lead nitrate, 100 μmol/kg). The effect of the two different proliferative stimuli on the appearance of γ -glutamyl transferase-positive foci was monitored by killing the rats for examination at 1, 2, 3, 5, and 6 days after the induction of cell proliferation. The results indicate that while enzyme-altered hepatocytes can be observed as early as 3 days after partial hepatectomy and are characterized by a rapid growth, direct hyperplasia did not exert any effect on the growth capacity of initiated cells. No effect of lead nitrate-induced hyperplasia was observed following three administrations of the mitogen. When platelet-poor plasma taken from animals exposed to the different proliferative stimuli was tested in primary cultures of hepatocytes, it was found that it induced a significant increase in the labeling index of normal hepatocytes. However, while serum taken 6 days after partial hepatectomy was still able to induce a significant increase in the labeling index, platelet-poor plasma from lead-treated rats had lost part of its effect at 5 days after treatment. The inability of direct hyperplasia to stimulate the development of enzyme-altered hepatic foci was not unique to lead nitrate since the same phenomenon was observed when three other hepatomitogens, nafenopin, cyproterone acetate, and ethylene dibromide, were used.     

Effects of water deprivation and angiotensin II intracerebroventricular administration on brain nitric oxide synthase activity

Intracranial administration of -arginine causes a reduction of the water intake induced by water deprivation or by intracerebroventricular (i.c.v.) injection of angiotensin II (angiotensin II), through the release of nitric oxide (NO) in the central nervous system. We studied the effects of i.c.v. angiotensin II (120 ng/rat) in association with i.c.v. -arginine (2.5–10 μg/rat) on blood pressure. We also studied the effects of both peripheral and central angiotensin II injection (1.5–6 mg kg⁻¹ i.p. and 30–120 ng rat⁻¹ i.c.v., respectively) on NO synthase activity in the cortex, diencephalon and brainstem, after water deprivation (24 h), conditions producing activation of the renin-angiotensin system. -arginine dose dependently antagonized the increase in blood pressure induced by i.c.v. angiotensin II (P<0.001). Peripheral administration of angiotensin II produced a dose-dependent reduction of NO synthase activity in the brainstem and cortex (P<0.001), but not in the diencephalon. Water deprivation produced similar effects on brain NO synthase activity. Angiotensin II i.c.v. injection caused NO synthase activity reduction in all brain regions studied (P<0.001). Our findings suggest that NO and angiotensin II could play opposite roles in brain regulation of blood pressure and drinking behaviour.