BACKGROUND

Fibroblast growth factor 23 (FGF-23) is important in the regulation of phosphorus and vitamin D metabolism. States of excess circulating FGF-23 are associated with renal phosphate wasting and inappropriately low serum <em>1</em>,25-dihydroxyvitamin D [<em>1</em>,25(OH)(2)D] concentrations. Conversely, states of absent or biologically inactive circulating FGF-23 are associated with increased serum phosphorus and <em>1</em>,25(OH)(2)D concentrations. Restriction of the dietary intake of phosphorus increases renal phosphate reabsorption and <em>1</em>,25(OH)(2)D production, whereas the opposite occurs when dietary phosphorus is supplemented.

OBJECTIVE

We sought to determine whether serum FGF-23 concentration is regulated by dietary phosphorus and thereby mediates the physiological response of serum <em>1</em>,25(OH)(2)D to changes in dietary phosphorus.

METHODS

We studied <em>1</em>3 healthy men as inpatients during a 4-wk dietary phosphorus intervention study.

METHODS

Subjects consumed a constant diet that provided 500 mg of phosphorus per day, which was supplemented to achieve three phosphorus intakes, each of 9 d: <em>1</em>) control = <em>1</em>500 mg/d; 2) supplemented = 2300 mg/d; 3) restricted = 625 mg/d. Intakes of calcium, sodium, potassium, magnesium, and energy were constant.

METHODS

Serum FGF-23, <em>1</em>,25(OH)(2)D, phosphorus, and calcium concentrations were measured.

RESULTS

Serum FGF-23 concentrations decreased significantly from 30.7 +/- 8.7 pg/ml during phosphorus supplementation to <em>1</em>9.6 +/- 7.0 pg/ml during phosphorus restriction. Serum <em>1</em>,25(OH)(2)D concentrations increased significantly from 29 +/- <em>1</em>0 pg/ml (75 +/- 26 pmol/liter) during phosphorus supplementation to 40 +/- <em>1</em>6 pg/ml (<em>1</em>04 +/- 42 pmol/liter) during phosphorus restriction (P < 0.00<em>1</em>). Serum <em>1</em>,25(OH)(2)D concentrations varied inversely with those of serum FGF-23 (r = -0.67, P < 0.00<em>1</em>).

CONCLUSIONS

We conclude that in healthy men, changes in dietary phosphorus within the physiological range of intakes regulate serum FGF-23 concentrations and suggest that dietary phosphorus regulation of <em>1</em>,25(OH)(2)D production is mediated, at least in part, by changes in circulating FGF-23.

OBJECTIVE

The aim of this study was to assess the effects of regular physical exercise on local inflammatory parameters in the skeletal muscle of patients with chronic heart failure (CHF).

BACKGROUND

Inflammatory activation with increased serum cytokine levels and expression of inducible nitric oxide synthase (iNOS) in the myocardium and peripheral skeletal muscles has been described in CHF.

METHODS

Twenty male patients with stable CHF (left ventricular ejection fraction 25 +/- 2%; age 54 +/- 2 years) were randomized to a training group (n = <em>1</em>0) or a control group (n = <em>1</em>0). At baseline and after six months, serum samples and vastus lateralis muscle biopsies were obtained. Serum tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, and IL-<em>1</em>-beta levels were measured by enzyme-linked immunosorbent assay, local cytokine, and iNOS expression by real-time polymerase chain reaction.

RESULTS

Exercise training improved peak oxygen uptake by 29% in the training group (from 20.3 +/- <em>1</em>.0 to 26.<em>1</em> +/- <em>1</em>.5 ml/kg. min; p < 0.00<em>1</em> vs. control group). While serum levels of TNF-alpha, IL-6, and IL-<em>1</em>-beta remained unaffected by training, local skeletal muscle TNF-alpha decreased from <em>1</em>.9 +/- 0.4 to <em>1</em>.2 +/- 0.3 relative U (p < 0.05 for change vs. control group), IL-6 from 7<em>1</em>.3 +/- <em>1</em>6.5 to 4<em>1</em>.3 +/- 8.8 relative U (p < 0.05 vs. begin), and IL-<em>1</em>-beta from 2.7 +/- <em>1</em>.<em>1</em> to <em>1</em>.4 +/- 0.6 relative U (p = 0.02 vs. control group). Exercise training also reduced local iNOS expression by 52% (from 6.3 +/- <em>1</em>.2 to 3.0 +/- <em>1</em>.0 relative U; p = 0.007 vs. control group).

CONCLUSIONS

Exercise training significantly reduced the local expression of TNF-alpha, IL-<em>1</em>-beta, IL-6, and iNOS in the skeletal muscle of CHF patients. These local anti-inflammatory effects of exercise may attenuate the catabolic wasting process associated with the progression of CHF.

OBJECTIVE

The purpose of this study was to develop a risk stratification score to predict warfarin-associated hemorrhage.

BACKGROUND

Optimal decision making regarding warfarin use for atrial fibrillation requires estimation of hemorrhage risk.

METHODS

We followed up 9,<em>1</em>86 patients with atrial fibrillation contributing 32,888 person-years of follow-up on warfarin, obtaining data from clinical databases and validating hemorrhage events using medical record review. We used Cox regression models to develop a hemorrhage risk stratification score, selecting candidate variables using bootstrapping approaches. The final model was internally validated by split-sample testing and compared with 6 published hemorrhage risk schemes.

RESULTS

We observed 46<em>1</em> first major hemorrhages during follow-up (<em>1</em>.4% annually). Five independent variables were included in the final model and weighted by regression coefficients: anemia (3 points), severe renal disease (e.g., glomerular filtration rate <30 <em>ml</em>/min or dialysis-dependent, 3 points), age ≥75 years (2 points), prior bleeding (<em>1</em> point), and hypertension (<em>1</em> point). Major hemorrhage rates ranged from 0.4% (0 points) to <em>1</em>7.3% per year (<em>1</em>0 points). Collapsed into a 3-category risk score, major hemorrhage rates were 0.8% for low risk (0 to 3 points), 2.6% for intermediate risk (4 points), and 5.8% for high risk (5 to <em>1</em>0 points). The c-index for the continuous risk score was 0.74 and 0.69 for the 3-category score, higher than in the other risk schemes. There was net reclassification improvement versus all 6 comparators (from 27% to 56%).

CONCLUSIONS

A simple 5-variable risk score was effective in quantifying the risk of warfarin-associated hemorrhage in a large community-based cohort of patients with atrial fibrillation.

BACKGROUND

The efficacy of highly active antiretroviral therapy (HAART) in suppression of HIV-<em>1</em> is well documented. We investigated virological and clinical outcomes of HAART in routine practice.

METHODS

We analysed prospective data from the Swiss HIV Cohort Study on suppression of viral load and progression to AIDS or death in 2674 outpatients (median age 36 years, 27.3% women) who started HAART in <em>1</em>995-98. Viral rebound was defined as two consecutive HIV-<em>1</em>-RNA measurements of more than 400 copies/mL. We analysed separately outcomes in patients with a history of antiretroviral treatment and in treatment-naïve patients.

RESULTS

An estimated 90.7% of treatment-naïve patients reached undetectable viral load (<400 copies/mL) by <em>1</em>2 months. Among pretreated patients, estimates ranged from 70.3% treated with one new drug to 78.7% on three new drugs. 2 years after reaching undetectable concentrations, an estimated 20.<em>1</em>% of treatment-naïve patients and 35.7-40.<em>1</em>% of pretreated patients had viral rebound. At 30 months, an estimated 6.6% (95% CI 4.6-8.6) of patients who had maintained undetectable concentrations, 9.0% (5.5-<em>1</em>2.5) who had viral rebound, and 20.<em>1</em>% (<em>1</em>5.3-24.9) who had never reached undetectable concentrations developed AIDS or died. Compared with patients who maintained undetectable viral load, the adjusted relative hazard of AIDS or death was <em>1</em>.00 (0.66-<em>1</em>.55) for patients with viral rebound, and 2.40 (<em>1</em>.72-3.33) for patients who failed to reach undetectable concentrations.

CONCLUSIONS

The rate of virological failure of HAART was high among these patients, but the probability of clinical progression was low even in patients with viral rebound.

BACKGROUND

This randomized phase III study was to evaluate the efficacy and safety of cytoreductive surgery (CRS) plus hyperthermic intraperitoneal chemotherapy (HIPEC) for the treatment of peritoneal carcinomatosis (PC) from gastric cancer.

METHODS

Sixty-eight gastric PC patients were randomized into CRS alone (n = 34) or CRS + HIPEC (n = 34) receiving cisplatin 120 mg and mitomycin C 30 mg each in 6000 ml of normal saline at 43 ± 0.5°C for 60-90 min. The primary end point was overall survival, and the secondary end points were safety profiles.

RESULTS

Major clinicopathological characteristics were balanced between the 2 groups. The PC index was 2-36 (median 15) in the CRS + HIPEC and 3-23 (median 15) in the CRS groups (P = 0.489). The completeness of CRS score (CC 0-1) was 58.8% (20 of 34) in the CRS and 58.8% (20 of 34) in the CRS + HIPEC groups (P = 1.000). At a median follow-up of 32 months (7.5-83.5 months), death occurred in 33 of 34 (97.1%) cases in the CRS group and 29 of 34 (85.3%) cases of the CRS + HIPEC group. The median survival was 6.5 months (95% confidence interval 4.8-8.2 months) in CRS and 11.0 months (95% confidence interval 10.0-11.9 months) in the CRS + HIPEC groups (P = 0.046). Four patients (11.7%) in the CRS group and 5 (14.7%) patients in the CRS + HIPEC group developed serious adverse events (P = 0.839). Multivariate analysis found CRS + HIPEC, synchronous PC, CC 0-1, systemic chemotherapy ≥ 6 cycles, and no serious adverse events were independent predictors for better survival.

CONCLUSIONS

For synchronous gastric PC, CRS + HIPEC with mitomycin C 30 mg and cisplatin 120 mg may improve survival with acceptable morbidity.

Chemoembolization with lipiodol (iodized oil) is widely used to treat patients with unresectable hepatocellular carcinoma. Severe side effects have been reported, and improved survival has not been clearly demonstrated.

Patients with unresectable hepatocellular carcinoma who did not have severe liver disease and who met additional entry criteria were randomly assigned to receive either lipiodol chemoembolization (70 mg of cisplatin, 10 ml of lipiodol, and gelatin-sponge [Gelfoam] particles delivered through the hepatic artery) or conservative management involving treatment of complications and pain. Courses of treatment were to be given every two months for a maximum of four courses. The main end point was survival.

The study was stopped in December, 1992, after a sequential analysis showed the lack of the expected benefit from chemoembolization. As of October 1, 1994, 39 of the 50 patients assigned to chemoembolization and 40 of the 46 patients assigned to conservative management had died. Twenty-six patients assigned to chemoembolization received all four courses of treatment. There was no significant difference in survival between the two groups, although there was a trend favoring the chemoembolization group (estimated relative risk of death in the control group, 1.4; 95 percent confidence interval, 0.9 to 2.2; P = 0.13). The comparison of survival between the two groups was not substantially changed by adjustments for differences in base-line and prognostic characteristics (adjusted relative risk, 1.3; 95 percent confidence interval, 0.8 to 2.1; P = 0.31). At one year, the estimated survival rates were 62 percent in the chemoembolization group (95 percent confidence interval, 48.6 to 75.4 percent) and 43.5 percent in the conservative-management group (95 percent confidence interval, 29.2 to 57.8 percent). In the chemoembolization group, tumor growth, as assessed by tumor size and serum alpha-fetoprotein concentration, was reduced and the incidence of portal obstruction was lower than in the conservative-management group. Liver failure occurred after 47 courses of treatment in 30 patients assigned to chemoembolization.

In a group of patients with unresectable hepatocellular carcinoma but without severe liver disease, lipiodol chemoembolization reduced tumor growth, often caused acute liver failure, and did not significantly improve survival.

BACKGROUND

Previous studies using autologous bone marrow mononuclear cells (BMCs) in patients with ischemic cardiomyopathy have demonstrated safety and suggested efficacy.

OBJECTIVE

To determine if administration of BMCs through transendocardial injections improves myocardial perfusion, reduces left ventricular end-systolic volume (LVESV), or enhances maximal oxygen consumption in patients with coronary artery disease or LV dysfunction, and limiting heart failure or angina.

METHODS

A phase 2 randomized double-blind, placebo-controlled trial of symptomatic patients (New York Heart Association classification II-III or Canadian Cardiovascular Society classification II-IV) with a left ventricular ejection fraction of 45% or less, a perfusion defect by single-photon emission tomography (SPECT), and coronary artery disease not amenable to revascularization who were receiving maximal medical therapy at 5 National Heart, Lung, and Blood Institute-sponsored Cardiovascular Cell Therapy Research Network (CCTRN) sites between April 29, 2009, and April <em>1</em>8, 20<em>1</em><em>1</em>.

METHODS

Bone marrow aspiration (isolation of BMCs using a standardized automated system performed locally) and transendocardial injection of <em>1</em>00 million BMCs or placebo (ratio of 2 for BMC group to <em>1</em> for placebo group).

METHODS

Co-primary end points assessed at 6 months: changes in LVESV assessed by echocardiography, maximal oxygen consumption, and reversibility on SPECT. Phenotypic and functional analyses of the cell product were performed by the CCTRN biorepository core laboratory.

RESULTS

Of <em>1</em>53 patients who provided consent, a total of 92 (82 men; average age: 63 years) were randomized (n = 6<em>1</em> in BMC group and n = 3<em>1</em> in placebo group). Changes in LVESV index (-0.9 mL/m(2) [95% CI, -6.<em>1</em> to 4.3]; P = .73), maximal oxygen consumption (<em>1</em>.0 [95% CI, -0.42 to 2.34]; P = .<em>1</em>7), and reversible defect (-<em>1</em>.2 [95% CI, -<em>1</em>2.50 to <em>1</em>0.<em>1</em>2]; P = .84) were not statistically significant. There were no differences found in any of the secondary outcomes, including percent myocardial defect, total defect size, fixed defect size, regional wall motion, and clinical improvement.

CONCLUSIONS

Among patients with chronic ischemic heart failure, transendocardial injection of autologous BMCs compared with placebo did not improve LVESV, maximal oxygen consumption, or reversibility on SPECT.

BACKGROUND

clinicaltrials.gov Identifier: NCT00824005.

Rare human immunodeficiency virus <em>1</em>-infected individuals, termed elite suppressors (ES), maintain plasma virus levels of <50 copies/<em>ml</em> and normal CD4 counts without therapy. The major histocompatibility complex class I allele group human histocompatibility leukocyte antigen (HLA)-B*57 is overrepresented in this population. Mutations in HLA-B*57-restricted epitopes have been observed in ES, but their significance has remained unclear. Here we investigate the extent and impact of cytotoxic T lymphocyte (CTL) escape mutations in HLA-B*57+ ES. We provide the first direct evidence that most ES experience chronic low level viremia. Sequencing revealed a striking discordance between the genotypes of plasma virus and archived provirus in resting CD4+ T cells. Mutations in HLA-B*57-restricted Gag epitopes were present in all viruses from plasma but were rare in proviruses, suggesting powerful selective pressure acting at these epitopes. Surprisingly, strong CD8+ T cell interferon-gamma responses were detected against some mutant epitopes found in plasma virus, suggesting the development of de novo responses to viral variants. In some individuals, relative CD8+ T cell interleukin-2 responses showed better correlation with the selection observed in vivo. Thus, analysis of low level viremia reveals an unexpectedly high level of CTL escape mutations reflecting selective pressure acting at HLA-B*57-restricted epitopes in ES. Continued viral suppression probably reflects CTL responses against unmutated epitopes and residual or de novo responses against epitopes with escape mutations.

BACKGROUND

Raltegravir is an HIV-<em>1</em> integrase strand-transfer inhibitor with potent in vitro activity. This study explored the antiretroviral activity and safety of raltegravir in treatment-naive patients with plasma HIV-<em>1</em> RNA levels>> or = 5000 copies/<em>mL</em> and CD4 T-cell counts>> or = <em>1</em>00 cells/mm.

METHODS

Multicenter, double-blind, randomized, controlled study of raltegravir at doses of <em>1</em>00, 200, 400, and 600 mg twice daily versus efavirenz at a dose of 600 mg/d, all in combination with tenofovir at a dose of 300 mg/d and lamivudine at a dose of 300 mg/d (clinicaltrials.gov identifier: NCT00<em>1</em>00048).

RESULTS

In the <em>1</em>98 patients treated (<em>1</em>60 on raltegravir and 38 on efavirenz), the mean HIV-<em>1</em> RNA level ranged from 4.6 to 4.8 log<em>1</em>0 copies/<em>mL</em> at baseline. At weeks 2, 4, and 8, the proportion of patients achieving an HIV-<em>1</em> RNA level <50 copies/<em>mL</em> was greater in each of the raltegravir treatment groups than in the efavirenz group. By week 24, all treatment groups appeared similar, with plasma HIV-<em>1</em> RNA levels <400 copies/<em>mL</em> in 85% to 98% of patients and <50 copies/<em>mL</em> in 85% to 95% of patients. These reductions were maintained through week 48 in 85% to 98% of patients and in 83% to 88% of patients, respectively. Five (3%) patients on raltegravir and <em>1</em> (3%) on efavirenz experienced virologic failure before week 48. Drug-related clinical adverse events were less common with raltegravir than with efavirenz. After 24 and 48 weeks of treatment, raltegravir did not result in increased serum levels of total cholesterol, low-density lipoprotein cholesterol, or triglycerides.

CONCLUSIONS

Raltegravir at all doses studied was generally well tolerated in combination with tenofovir and lamivudine. Raltegravir exhibited potent and durable antiretroviral activity similar to that of efavirenz at 24 and 48 weeks but achieved HIV-<em>1</em> RNA levels below detection at a more rapid rate.

The present study examined the roles of peroxisome proliferator-activated receptors (PPAR) in activation of hepatic stellate cells (HSC), a pivotal event in liver fibrogenesis. RNase protection assay detected mRNA for PPARgamma<em>1</em> but not that for the adipocyte-specific gamma2 isoform in HSC isolated from sham-operated rats, whereas the transcripts for neither isoforms were detectable in HSC from cholestatic liver fibrosis induced by bile duct ligation (BDL). Semi-quantitative reverse transcriptase-polymerase chain reaction confirmed a 70% reduction in PPARgamma mRNA level in HSC from BDL. Nuclear extracts from BDL cells showed an expected diminution of binding to PPAR-responsive element, whereas NF-kappaB and AP-<em>1</em> binding were increased. Treatment of cultured-activated HSC with ligands for PPARgamma (<em>1</em>0 microm <em>1</em>5-deoxy-Delta(<em>1</em>2,<em>1</em>4)-PGJ(2) (<em>1</em>5dPGJ(2)); 0.<em>1</em> approximately <em>1</em>0 microm BRL49653) inhibited DNA and collagen synthesis without affecting the cell viability. Suppression of HSC collagen by <em>1</em>5dPGJ(2) was abrogated 70% by the concomitant treatment with a PPARgamma antagonist (GW9662). HSC DNA and collagen synthesis were inhibited by WY<em>1</em>4643 at the concentrations known to activate both PPARalpha and gamma >><em>1</em>00 microm) but not at those that only activate PPARalpha ((<em>1</em>0 microm) or by a synthetic PPARalpha-selective agonist (GW9578). <em>1</em>5dPGJ(2) reduced alpha<em>1</em>(I) procollagen, smooth muscle alpha-actin, and monocyte chemotactic protein-<em>1</em> mRNA levels while inducing matrix metalloproteinase-3 and CD36. <em>1</em>5dPGJ(2) and BRL49653 inhibited alpha<em>1</em>(I) procollagen promoter activity. Tumor necrosis factor alpha (<em>1</em>0 ng/<em>ml</em>) reduced PPARgamma mRNA, and this effect was prevented by the treatment with <em>1</em>5dPGJ(2). These results demonstrate that HSC activation is associated with the reductions in PPARgamma expression and PPAR-responsive element binding in vivo and is reversed by the treatment with PPARgamma ligands in vitro. These findings implicate diminished PPARgamma signaling in molecular mechanisms underlying activation of HSC in liver fibrogenesis and the potential therapeutic value of PPARgamma ligands for liver fibrosis.

BACKGROUND

Continuous passive motion after major knee surgery optimizes the functional prognosis but causes severe pain. The authors tested the hypothesis that postoperative analgesic techniques influence surgical outcome and the duration of convalescence.

METHODS

Before standardized general anesthesia, 56 adult scheduled for major knee surgery were rando<em>ml</em>y assigned to one of three groups, each to receive a different postoperative analgesic technique for 72 h: continuous epidural infusion, continuous femoral block, or intravenous patient-controlled morphine (dose, <em>1</em> mg; lockout interval, 7 min; maximum dose, 30 mg/4 h). The first two techniques were performed using a solution of <em>1</em>% lidocaine, 0.03 mg/<em>ml</em> morphine, and 2 microg/<em>ml</em> clonidine administered at 0.<em>1</em> <em>ml</em> x kg(-<em>1</em>) x h(-<em>1</em>). Pain was assessed at rest and during continuous passive motion using a visual analog scale. The early postoperative maximal amplitude of knee flexion was measured during continuous passive motion at 24 h and 48 h and compared with the target levels prescribed by the surgeon. To evaluate functional outcome, the maximal amplitudes were measured again on postoperative day 5, at hospital discharge (day 7), and at <em>1</em>- and 3-month follow-up examinations. When the patients left the surgical ward, they were admitted to a rehabilitation center, where their length of stay depended on prospectively determined discharge criteria

RESULTS

The continuous epidural infusion and continuous femoral block groups showed significantly lower visual analog scale scores at rest and during continuous passive motion compared with the patient-controlled morphine group. The early postoperative knee mobilization levels in both continuous epidural infusion and continuous femoral block groups were significantly closer to the target levels prescribed by the surgeon than in the patient-controlled morphine group. On postoperative day 7, these values were 90 degrees (60-<em>1</em>00 degrees)(median and 25th-75th percentiles) in the continuous epidural infusion group, 90 degrees (60-<em>1</em>00 degrees) in the continuous femoral block group, and 80 degrees (60-<em>1</em>00 degrees) in the patient-controlled morphine group (P < 0.05). The durations of stay in the rehabilitation center were significantly shorter: 37 days (range, 30-45 days) in the continuous epidural infusion group, 40 days (range, 3<em>1</em>-60 days) in the continuous femoral block group, and 50 days (range, 30-80 days) in the patient-controlled morphine group (P < 0.05). Side effects were encountered more frequently in the continuous epidural infusion group.

CONCLUSIONS

Regional analgesic techniques improve early rehabilitation after major knee surgery by effectively controlling pain during continuous passive motion, thereby hastening convalescence.

Patients in whom virologic suppression is achieved with highly active antiretroviral therapy (HAART) retain long-lived cellular reservoirs of human immunodeficiency virus type <em>1</em> (HIV-<em>1</em>); this retention is an obstacle to sustained control of infection. To assess the impact that initiating treatment during primary HIV-<em>1</em> infection has on this cell population, we analyzed the decay kinetics of HIV-<em>1</em> DNA and of infectivity associated with cells activated ex vivo in 27 patients who initiated therapy before or <6 months after seroconversion and in whom viremia was suppressed to <50 copies/<em>mL</em>. The clearance rates of cellular reservoirs could not be distinguished by these techniques (median half-life, 20 weeks) during the first year of HAART. The clearance of HIV-<em>1</em> DNA slowed significantly during the subsequent 3 years of treatment (median half-life, 70 weeks), consistent with heterogeneous cellular reservoirs being present. Total cell-associated infectivity (CAI) after <em>1</em> year of treatment was undetectable (<0.07 infectious units/million cells [IUPM]) in most patients initiating treatment during primary infection either before (9/9) or <6 months after (6/8) seroconversion. In contrast, all <em>1</em>7 control patients who initiated HAART during chronic infection retained detectable CAI after 3-6 years of treatment (median reservoir size, <em>1</em>.<em>1</em> IUPM; P<.0005). These results suggest that treatment <6 months after seroconversion may facilitate long-term control of cellular reservoirs that maintain HIV-<em>1</em> infection during treatment.

Pomegranate (Punica granatum L.) fruits are widely consumed as juice (PJ). The potent antioxidant and anti-atherosclerotic activities of PJ are attributed to its polyphenols including punicalagin, the major fruit ellagitannin, and ellagic acid (EA). Punicalagin is the major antioxidant polyphenol ingredient in PJ. Punicalagin, EA, a standardized total pomegranate tannin (TPT) extract and PJ were evaluated for in vitro antiproliferative, apoptotic and antioxidant activities. Punicalagin, EA and TPT were evaluated for antiproliferative activity at <em>1</em>2.5-<em>1</em>00 microg/<em>ml</em> on human oral (KB, CAL27), colon (HT-29, HCT<em>1</em><em>1</em>6, SW480, SW620) and prostate (RWPE-<em>1</em>, 22Rv<em>1</em>) tumor cells. Punicalagin, EA and TPT were evaluated at <em>1</em>00 microg/<em>ml</em> concentrations for apoptotic effects and at <em>1</em>0 microg/<em>ml</em> concentrations for antioxidant properties. However, to evaluate the synergistic and/or additive contributions from other PJ phytochemicals, PJ was tested at concentrations normalized to deliver equivalent amounts of punicalagin (w/w). Apoptotic effects were evaluated against the HT-29 and HCT<em>1</em><em>1</em>6 colon cancer cell lines. Antioxidant effects were evaluated using inhibition of lipid peroxidation and Trolox equivalent antioxidant capacity (TEAC) assays. Pomegranate juice showed greatest antiproliferative activity against all cell lines by inhibiting proliferation from 30% to <em>1</em>00%. At <em>1</em>00 microg/<em>ml</em>, PJ, EA, punicalagin and TPT induced apoptosis in HT-29 colon cells. However, in the HCT<em>1</em><em>1</em>6 colon cells, EA, punicalagin and TPT but not PJ induced apoptosis. The trend in antioxidant activity was PJ>TPT>punicalagin>EA. The superior bioactivity of PJ compared to its purified polyphenols illustrated the multifactorial effects and chemical synergy of the action of multiple compounds compared to single purified active ingredients.

OBJECTIVE

This study was aimed at detecting and characterizing circulating tumor cells (CTC) before and after neoadjuvant therapy (NT) in the peripheral blood of patients with breast cancer.

METHODS

The clinical trial GeparQuattro incorporated NT approaches (epirubicin/cyclophosphamide prior to randomization to docetaxel alone, docetaxel in combination with capecitabine, or docetaxel followed by capecitabine) and additional trastuzumab treatment for patients with HER2-positive tumors. We used the Food and Drug Administration-approved CellSearch system for CTC detection and evaluation of HER2 expression and developed HER2 immunoscoring for CTC.

RESULTS

We detected>> or =<em>1</em> CTC/7.5 <em>mL</em> in 46 of 2<em>1</em>3 patients (2<em>1</em>.6%) before NT and in 22 of 207 patients (<em>1</em>0.6%) after NT (P = 0.002). Twenty (<em>1</em>5.0%) initially CTC-positive cases were CTC-negative after NT, whereas <em>1</em><em>1</em> (8.3%) cases were CTC-positive after NT, although no CTC could be found before NT. CTC detection did not correlate with primary tumor characteristics. Furthermore, there was no association between tumor response to NT and CTC detection. HER2-overexpressing CTC were observed in <em>1</em>4 of 58 CTC-positive patients (24.<em>1</em>%), including 8 patients with HER2-negative primary tumors and 3 patients after trastuzumab treatment. CTC scored HER2-negative or weakly HER2-positive before or after NT were present in <em>1</em><em>1</em> of 2<em>1</em> patients with HER2-positive primary tumors. HER2 overexpression on CTC was restricted to ductal carcinomas and associated with high tumor stage (P = 0.002).

CONCLUSIONS

CTC number was low in patients with primary breast cancer. The decrease in CTC incidence during treatment was not correlated with standard clinical characteristics and primary tumor response. Information on the HER2 status of CTC might be helpful for stratification and monitoring of HER2-directed therapies.

Normal peripheral blood mononuclear cells demonstrated increased DNA synthesis and secretion of newly synthesized protein when suboptimal concentrations of Concanavalin A (Con A) were added to the cultures after 24-h incubation in vitro. Cells stimulated by Con A, <em>1</em> mug/<em>ml</em>, after 24-h incubation demonstrated 3.0 times more tritiated thymidine incorporation, and 4.4 times more <em>1</em>4C-amino acid incorporation into newly synthesized secreted protein, than cells stimulated at 0 h (P less than 0.00<em>1</em>). The acquisition of increased responsiveness was not abrogated by washing and resuspending the cells in fresh medium. Since the increased responsiveness could be inhibited by the addition to the cultures of small numbers of cells previously activated by Con A it is suggested that the enhanced reactivity acquired in culture represents the loss of a subpopulation of suppressor cells that modulate the T-lymphocyte response. Cells from nine patients with active, untreated systemic lupus erythematosus demonstrated normal responses to optimal concentrations of Con A added at 0 h, but an impaired response to Con A, <em>1</em> mug/<em>ml</em>. When these cells were incubated for 24 h, a significant increased response to Con A was not observed. This observation suggests that patients with active SLE lack circulating suppressor cells. When seven SLE patients were again studied after corticosteroid therapy had led to clinical improvement, the response to Con A, <em>1</em> mug/<em>ml</em>, added after 24-h incubation was similar to that observed in normal controls, suggesting that suppressor function in SLE returns as disease activity declines.

The site of action of the immunosuppressive drug cyclosporin A in in vitro cytotoxic allograft responses has been localized. General cytotoxic effects of the drug on proliferating T cells became apparent at concentrations of 500-<em>1</em>000 ng/<em>ml</em>, while selective effects were observed at concentrations of <em>1</em>0-<em>1</em>00 ng/<em>ml</em>. The selective effects included a blockade of interleukin 2 release from activated T helper cells on the one hand and inhibition of interleukin <em>1</em> release from splenic adherent cells on the other. While cyclosporin A did not interfere with the intracellular events required for the activation and subsequent clonal expansion of alloreactive T cells, the lack of interleukin <em>1</em> and interleukin 2 induced by cyclosporin A results in an inability of T responder cells to mount cytotoxic allograft responses in vitro.

BACKGROUND

The 2NN Study was a randomised comparison of the non-nucleoside reverse-transcriptase inhibitors (NNRTI) nevirapine and efavirenz.

METHODS

In this multicentre, open-label, randomised trial, <em>1</em>2<em>1</em>6 antiretroviral-therapy-naive patients were assigned nevirapine 400 mg once daily, nevirapine 200 mg twice daily, efavirenz 600 mg once daily, or nevirapine (400 mg) and efavirenz (800 mg) once daily, plus stavudine and lamivudine, for 48 weeks. The primary endpoint was the proportion of patients with treatment failure (less than <em>1</em> log(<em>1</em>0) decline in plasma HIV-<em>1</em> RNA in the first <em>1</em>2 weeks or two consecutive measurements of more than 50 copies per <em>mL</em> from week 24 onwards, disease progression [new Centers for Disease Control and Prevention grade C event or death], or change of allocated treatment). Analyses were by intention to treat.

RESULTS

Treatment failure occurred in 96 (43.6%) of 220 patients assigned nevirapine once daily, <em>1</em>69 (43.7%) of 387 assigned nevirapine twice daily, <em>1</em>5<em>1</em> (37.8%) of 400 assigned efavirenz, and <em>1</em><em>1</em><em>1</em> (53.<em>1</em>%) of 209 assigned nevirapine plus efavirenz. The difference between nevirapine twice daily and efavirenz was 5.9% (95% CI -0.9 to <em>1</em>2.8). There were no significant differences among the study groups in the proportions with plasma HIV-<em>1</em> RNA concentrations below 50 copies per <em>mL</em> at week 48 (p=0.<em>1</em>93) or the increases in CD4-positive cells (p=0.800). Nevirapine plus efavirenz was associated with the highest frequency of clinical adverse events, and nevirapine once daily with significantly more hepatobiliary laboratory toxicities than efavirenz. Of 25 observed deaths, two were attributed to nevirapine.

CONCLUSIONS

Antiretroviral therapy with nevirapine or efavirenz showed similar efficacy, so triple-drug regimens with either NNRTI are valid for first-line treatment. There are, however, differences in safety profiles. Combination of nevirapine and efavirenz did not improve efficacy but caused more adverse events.

OBJECTIVE

The ItalKid Project is a prospective, population-based registry that was started in <em>1</em>990 with the aim of assessing the epidemiology of childhood chronic renal failure (CRF), describing the natural history of the disease, and identifying factors that influence its course. This article reports the epidemiologic results.

METHODS

Prevalent and incident cases of CRF in children and adolescents were identified throughout Italy (total population base: <em>1</em>6.8 million children) by regularly asking all of the pediatric hospitals and adult nephrology units potentially involved in caring for children with kidney disease to report all cases that meet the inclusion criteria and then to update the clinical information regarding all previously reported patients on an annual basis. The inclusion criteria were <em>1</em>) creatinine clearance (Ccr; according to Schwartz's formula) <75 <em>mL</em>/min/<em>1</em>.73 m2 bsa (predialysis) and 2) an age of <20 years at the time of registration.

RESULTS

By December 3<em>1</em>, 2000, <em>1</em><em>1</em>97 patients (803 boys) had been registered. The mean incidence was <em>1</em>2.<em>1</em> cases per million (range: 8.8-<em>1</em>3.9), and the (point) prevalence was 74.7 per million of the age-related population. The mean age at registration was 6.9 +/- 5.4 years, and the mean Ccr was 4<em>1</em>.7 +/- 20.5 <em>mL</em>/min/<em>1</em>.73 m2. The leading causes of CRF were hypodysplasia associated with urinary tract malformations (53.6%) and isolated hypodysplasia (<em>1</em>3.9%), whereas glomerular disease accounted for as few as 6.8%. Hypodysplasia associated with primary vesicoureteral reflux (VUR) alone was responsible for as many as 25.8% of the cases, thus being the leading single cause with a female-to-male ratio of <em>1</em>:3.2. The diagnosis of VUR was established early in life at an overall median age of 3 months (range: 0-<em>1</em>80). However, the diagnosis was made significantly later among girls, whose median age at diagnosis was 9 months (range: 0-<em>1</em>56; 95% confidence interval: 2<em>1</em>.2-49.3) as against 2 months among boys (range: 0-<em>1</em>80; 95% confidence interval: <em>1</em>0.9-2<em>1</em>.2). As many as 23.6% of the registered patients had at least <em>1</em> severe associated disease (excluding urological abnormalities). A steep decline in renal survival occurred during puberty and early postpuberty, leading almost 70% of the patients to end-stage renal failure by the age of 20 years. When the population was subdivided on the basis of Ccr at the time of registration, the probability of kidney survival at 20 years of age was significantly different, being 63% in patients with mild renal failure (Ccr 5<em>1</em>-75 <em>mL</em>/min), 30% in those with moderate renal failure (Ccr 25-50 <em>mL</em>/min), and 3% in those with severe renal failure (Ccr <25 <em>mL</em>/min). The incidence of renal replacement therapy was 7.3/y/<em>1</em>00 patients, and the case-fatality rate on conservative treatment was <em>1</em>.4<em>1</em>%.

CONCLUSIONS

This study provides important and recent epidemiologic information concerning CRF in children and adolescents: a mean annual incidence of <em>1</em>2.<em>1</em> new patients per million of the age-related population with a very high proportion (57.6%) of hypodysplastic renal diseases with or without urinary tract malformation. By the age of 20 years, the cumulative probability of end-stage renal disease in the population as a whole was 68%. The probability of kidney survival sharply declined during puberty and early postpuberty. This is the first prospective evaluation of the incidence and outcome of CRF in children, including those with mild and moderate renal impairment.

Amyloid was extracted from the spleen of a patient with primary amyloidosis by homogenizing it at high speed with water after preliminary treatments, first to remove proteins soluble in saline, and then to remove salts. The extracts containing amyloid appeared to be clear at concentrations up to 6 mg/<em>ml</em> of protein. The material gave little sediment on being centrifuged up to 20,000 g for <em>1</em> hr, but the protein was sedimented at <em>1</em>00,000 g in <em>1</em> hr. The amyloid could be precipitated from the extracts by addition of NaCl to 0.0075 mole/liter or of CaCl(2) to 0.0025 mole/liter. The protein-bound Congo red formed a red precipitate and this property was used to estimate recovery and purity of amyloid during extraction. On electronmicroscopy the isolated amyloid proved to be morphologically pure. It existed either as single filaments measuring 60-80 A in diameter or as large aggregates of these filaments.Freshly isolated amyloid in water sedimented as a single homogeneous peak with an s degrees (20,[unk]) of about 45-50S. On standing, the solution became cloudy and more rapidly sedimenting components appeared. On electrophoresis the material migrated as a homogeneous peak towards the anode. The protein had an amino acid composition different from that of all known serum proteins. It was rich in acidic amino acids and had little cysteine and methionine and no hydroxyproline. The total content of carbohydrate was less than 2%.

BACKGROUND

Acute renal dysfunction (ARD) and subsequent acute renal failure after cardiac surgery are associated with high mortality and morbidity. Early therapeutic or preventive intervention is hampered by the lack of an early biomarker for acute renal injury. Recent studies showed that urinary neutrophil gelatinase-associated lipocalin (NGAL or lipocalin 2) is up-regulated early (within <em>1</em>-3 h) after murine renal injury and in pediatric ARD after cardiac surgery. The authors hypothesized that postoperative urinary NGAL concentrations are increased in adult patients developing ARD after cardiac surgery compared with patients without ARD.

METHODS

After institutional review board approval, 8<em>1</em> cardiac surgical patients were prospectively studied. Urine samples were collected immediately before incision and at various time intervals after surgery for NGAL analysis by quantitative immunoblotting. ARD was defined as peak postoperative serum creatinine increase by 50% or greater compared with preoperative serum creatinine.

RESULTS

Sixteen of 8<em>1</em> patients (20%) developed postoperative ARD, and the mean urinary NGAL concentrations in patients who developed ARD were significantly higher early after surgery (after <em>1</em> h: 4,<em>1</em>95 +/- 6,520 [mean +/- SD] vs. <em>1</em>,068 +/- 2,<em>1</em>29 ng/ml; P < 0.0<em>1</em>) compared with patients who did not develop ARD. Mean urinary NGAL concentrations continued to increase and remained significantly higher at 3 and <em>1</em>8 h after cardiac surgery in patients with ARD. In contrast, urinary NGAL in patients without ARD decreased rapidly after cardiac surgery.

CONCLUSIONS

Patients developing postoperative ARD had significantly higher urinary NGAL concentrations early after cardiac surgery. Urinary NGAL may therefore be a useful early biomarker of ARD after cardiac surgery. These findings may facilitate the early detection of acute renal injury and potentially prevent progression to acute renal failure.

The chemokine receptors CCR2 and CCR5 play important roles in the recruitment of monocytes/macrophages and T cells. To better understand the role of both receptors in murine models of inflammatory diseases and to recognize potential problems when correlating these data to humans, we have generated mAbs against murine CCR2 and CCR5. In mice CCR2 is homogeneously expressed on monocytes and on 2--<em>1</em>5% of T cells, closely resembling the expression pattern in humans. In contrast to humans, murine NK cells are highly CCR5 positive. In addition, CCR5 is expressed on 3--<em>1</em>0% of CD4 and <em>1</em>0--40% of CD8-positive T cells and is weakly detectable on monocytes. Using a model of immune complex nephritis, we examined the effects of inflammation on chemokine receptor expression and found a <em>1</em>0-fold enrichment of CCR5(+) and CCR2(+) T cells in the inflamed kidneys. The activity of various chemokines and the antagonistic properties of the mAbs were measured by ligand-induced internalization of CCR2 and CCR5 on primary leukocytes. The Ab MC-2<em>1</em> (anti-CCR2) reduced the activity of murine monocyte chemotactic protein <em>1</em> by 95%, whereas the Ab MC-68 (anti-CCR5) blocked over 99% of the macrophage-inflammatory protein <em>1</em>alpha and RANTES activity. MC-2<em>1</em> and MC-68 efficiently blocked the ligand binding to CCR2 and CCR5 with an IC(50) of 0.09 and 0.6--<em>1</em>.0 microg/<em>ml</em>, respectively. In good correlation to these in vitro data, MC-2<em>1</em> almost completely prevented the influx of monocytes in thioglycollate-induced peritonitis. Therefore, both Abs appear as useful reagents to further study the role of CCR2 and CCR5 in murine disease models.

BACKGROUND

The biological mechanisms by which depression might increase risk of cardiovascular disease are not clear. Inflammation may be a key element in the development of atherosclerotic cardiovascular disease. Our objective was to determine the association between major depression and elevated C-reactive protein (CRP) level in a nationally representative cohort.

METHODS

We estimated the odds of elevated CRP level (>0.21 mg/mL) associated with depression in 6914 noninstitutionalized men and women (age, 18-39 years) from the Third National Health and Nutrition Examination Survey (NHANES III).

RESULTS

The prevalence of lifetime major depression was 5.7% for men and 11.7% for women. The prevalence of elevated CRP level was 13.7% for men and 27.3% for women. A history of major depression was associated with elevated CRP level (odds ratio [OR], 1.64; 95% confidence interval [CI], 1.20-2.24). The association between depression and CRP was much stronger among men than among women. Results were adjusted for age, African American race, body mass index, total cholesterol, log triglycerides, diabetes, systolic blood pressure, smoking status, alcohol use, estrogen use in women, aspirin use, ibuprofen use, and self-reported health status. Compared with men without a history of depression, CRP levels were higher among men who had a more recent (within 1 year) episode of depression (adjusted OR, 3.00; 95% CI, 1.39-6.48) and who had recurrent >>or=2 episodes) depression (adjusted OR, 3.55; 95% CI, 1.55-8.14).

CONCLUSIONS

Major depression is strongly associated with increased levels of CRP among men and could help explain the increased risk of cardiovascular disease associated with depression in men.

Following large-scale roll-out of antiretroviral therapy in sub-Saharan Africa, the non-clinical efficacy of antiretroviral therapy has received little attention. We aimed to systematically review virological efficacy and drug-resistance outcomes of programmes of antiretroviral therapy in sub-Saharan Africa. 89 studies with heterogeneous design, definitions, and methods were identified. Overall, in on-treatment analysis, <em>1</em>0 35<em>1</em> (78%) of <em>1</em>3 288 patients showed virological suppression after 6 months of antiretroviral therapy, 74<em>1</em>3 (76%) of 9794 after <em>1</em>2 months, and 3840 (67%) of 5690 after 24 months. Long-term virological data are scarce. Genotyping results were available for patients with virological failure (HIV-<em>1</em> RNA greater than <em>1</em>000 copies per <em>mL</em>). Most patients (839 of 849; 99%) were infected with a non-B HIV-<em>1</em> subtype. However, drug-resistance patterns were largely similar to those in subtype B. Resistance profiles were associated with the antiretroviral drugs commonly used: the lamivudine-associated M<em>1</em>84V mutation was most common, followed by K<em>1</em>03N which is associated with non-nucleoside reverse transcriptase inhibitors. Thymidine-analogue mutations and the K65R mutation were less common. First-line antiretroviral therapy regimens used in sub-Saharan Africa are effective. Profiles of drug resistance suggest that a second-line treatment regimen based on protease inhibitors, with a backbone of nucleoside reverse transcriptase inhibitors, is a reasonable option for patients with HIV in sub-Saharan Africa who experience first-line treatment failure.

Incubation for a short time of arachidonic acid with the microsomal fraction of a homogenate of the vesicular gland of sheep in the presence of <em>1</em> mM p-mercuribenzoate followed by extraction and silicic acid chromatography yielded two prostaglandin endoperoxides. The structures of these compounds, i.e., <em>1</em>5-hydroperoxy-9alpha,<em>1</em><em>1</em>alpha-peroxidoprosta-5,<em>1</em>3-dienoic acid (prostaglandin G(2)) and <em>1</em>5-hydroxy-9alpha,<em>1</em><em>1</em>alpha-peroxidoprosta-5,<em>1</em>3-dienoic acid (prostaglandin H(2)), were assigned mainly by a number of chemical transformations into previously known prostaglandins. The new prostaglandins were 50-200 times (prostaglandin G(2)) and <em>1</em>00-450 times (prostaglandin H(2)) more active than prostaglandin E(2) on the superfused aorta strip. The half-life of the prostaglandin endoperoxides in aqueous medium (about 5 min) was significantly longer than that of "rabbit aorta-contracting substance" released from guinea pig lung, indicating that none of the prostaglandin endoperoxides is identical with this factor. Addition of <em>1</em>0-300 ng/<em>ml</em> of the endoperoxides to suspensions of washed human platelets resulted in rapid aggregation. Furthermore, platelet aggregation induced by thrombin was accompanied by release of material reducible by stannous chloride into prostaglandin F(2alpha), thus indicating the involvement of endogenous prostaglandin endoperoxides in platelet aggregation.