Targeted cell replacement with bone marrow cells for airway epithelial regeneration

It has been suggested that some adult bone marrow cells (BMC) can localize to the lung and develop tissue-specific characteristics including those of pulmonary epithelial cells. Here, we show that the combination of mild airway injury (naphthalene-induced) as a conditioning regimen to direct the site of BMC localization and transtracheal delivery of short-term cultured BMC enhances airway localization and adoption of an epithelial-like phenotype. Confocal analysis of airway and alveolar-localized BMC (fluorescently labeled) with epithelial markers shows expression of the pulmonary epithelial proteins, Clara cell secretory protein, and surfactant protein C. To confirm epithelial gene expression by BMC, we generated transgenic mice expressing green fluorescent protein (GFP) driven by the epithelial-specific cytokeratin-18 promoter and injected BMC from these mice transtracheally into wild-type recipients after naphthalene-induced airway injury. BMC retention in the lung was observed for at least 120 days following cell delivery with increasing GFP transgene expression over time. Some BMC cultured in vitro over time also expressed GFP transgene, suggesting epithelial transdifferentiation of the BMC. The results indicate that targeted delivery of BMC can promote airway regeneration.

Mice with a targeted disruption of the Fgfrl1 gene die at birth due to alterations in the diaphragm

FGFRL1 is a recently discovered member of the fibroblast growth factor receptor family that is lacking the intracellular tyrosine kinase domain. To elucidate the function of the novel receptor, we created mice with a targeted disruption of the Fgfrl1 gene. These mice develop normally until term, but die within a few minutes after birth due to respiratory failure. The respiratory problems are explained by a significant reduction in the size of the diaphragm muscle, which is not sufficient to inflate the lungs after birth. The remaining portion of the diaphragm muscle appears to be well developed and innervated. It consists of differentiated myofibers with nuclei at the periphery. Fast and slow muscle fibers occur in normal proportions. The myogenic regulatory factors MyoD, Myf5, myogenin and Mrf4 and the myocyte enhancer factors Mef2A, Mef2B, Mef2C and Mef2D are expressed at normal levels. Experiments with a cell culture model involving C2C12 myoblasts show that Fgfrl1 is expressed during the late stages of myotube formation. Other skeletal muscles do not appear to be affected in the Fgfrl1 deficient mice. Thus, Fgfrl1 plays a critical role in the development of the diaphragm.

Aquatic therapies in patients with compromised left ventricular function and heart failure

With water immersion, gravity is partly eliminated, and the water exerts a pressure on the body surface. Consequently there is a blood volume shift from the periphery to the central circulation, resulting in marked volume loading of the thorax and heart. This paper presents a selection of published literature on water immersion, balneotherapy, aqua exercises, and swimming, in patients with left ventricular dysfunction (LVD) and/or stable chronic heart failure (CHF). Based on exploratory studies, central hemodynamic and neurohumoral responses of aquatic therapies will be illustrated. Major findings are: 1. In LVD and CHF, a positive effect of therapeutic warm-water tub bathing has been observed, which is assumed to be from afterload reduction due to peripheral vasodilatation caused by the warm water. 2. In coronary patients with LVD, at low-level water cycling the heart is working more efficiently than at lowlevel cycling outside of water. 3. In patients with previous extensive myocardial infarction, upright immersion to the neck resulted in temporary pathological increases in mean pulmonary artery pressure (mPAP) and mean pulmonary capillary pressures (mPCP). 4. Additionally, during slow swimming (20-25m/min) the mPAP and/or PCP were higher than during supine cycling outside water at a 100W load. 5. In CHF patients, neck- deep immersion resulted in a decrease or no change in stroke volume. 6. Although patients are hemodynamically compromised, they usually maintain a feeling of well-being during aquatic therapy. Based on these findings, clinical indications for aquatic therapies are proposed and ideas are presented to provoke further research.

Impact of body mass index on the one-year clinical outcome of patients undergoing multivessel revascularization with sirolimus-eluting stents (from the Arterial Revascularization Therapies Study Part II)

The differential safety and efficacy profiles of sirolimus-eluting stents when implanted in patients with multivessel coronary artery disease who have increased body mass indexes (BMIs) compared with those with normal BMIs are largely unknown. This study evaluated the impact of BMI on 1-year outcomes in patients with multivessel coronary artery disease treated with sirolimus-eluting stents as part of the Arterial Revascularization Therapies Study Part II (ARTS II). From February to November 2003, 607 patients were included at 45 centers; 176 patients had normal BMIs (<25 kg/m(2)), 289 were overweight (> or =25 and < or =30 kg/m(2)), and 142 were obese (>30 kg/m(2)). At 30 days, the cumulative incidence of the primary combined end point of death, myocardial infarction, cerebrovascular accident, and repeat revascularization (major adverse cardiac and cerebrovascular events) was 3.4% in the group with normal BMIs, 3.1% in overweight patients, and 2.8% in obese patients (p = 0.76). At 1 year, the cumulative incidence of major adverse cardiac and cerebrovascular events was 10.8%, 11.8%, and 7.0% in the normal BMI, overweight, and obese groups, respectively (p = 0.31). In conclusion, BMI had no impact on 1-year clinical outcomes in patients with multivessel coronary artery disease treated with sirolimus-eluting stents in ARTS II.

Early clinical trial experience with vaccine therapies in non-small-cell lung cancer

Cancer immunotherapy has made great progress because of advances in immunology and molecular biology. Increased understanding of mechanisms by which lung cancer cells escape the immune system and recognition of key tumor antigens and immune system components involved in tumor ignorance have led to the development of a variety of lung cancer vaccines. Immunotherapy has advanced from using nonspecific immunomodulatory agents to lung cancer-specific tumor antigens and tumor cell-derived vaccines. While understanding of immune processes and malignancy has improved, there is great opportunity for further research of vaccine therapies in non-small-cell lung cancer. Herein, we review the development and evolution of early lung cancer vaccine trials.

Dual induction of PKR with E2F-1 and IFN-alpha to enhance gene therapy against hepatocellular carcinoma

Overexpression of the transcription factor E2F-1 induces apoptosis in tumor cells. This apoptotic effect is partly mediated through the induction of the double-stranded RNA-activated protein kinase (PKR). Here, we investigate if agents that upregulate PKR could enhance the apoptotic effect of E2F-1 overexpression in liver tumors. In human hepatocellular carcinoma (HCC) cells (Hep3B, HepG2, Huh7), adenovirus-mediated overexpression of E2F-1 (AdCMV-E2F) transcriptionally increased PKR mRNA. The subsequent increase of total and phosphorylated PKR protein was followed by induction of apoptosis. When AdCMV-E2F was combined with the PKR modifier interferon alpha (IFNalpha), PKR was additionally upregulated and both PKR activation and apoptosis were increased. Subcutaneous xenograft tumors were selectively targeted using an adenoviral vector expressing E2F-1 under the control of the human telomerase reverse transcriptase (hTERT) promoter (AdhTERT-E2F). Weekly systemic administration of AdhTERT-E2F inhibited tumor growth. The tumor suppressive effect of AdhTERT-E2F therapy was further enhanced in combination with IFNalpha.Our results demonstrate that PKR activating agents enhance the anti-tumor effect of E2F-1 overexpression in HCC in-vitro and in-vivo. Hence, modulation of PKR is a potential strategy to increase the efficacy of PKR-dependent anti-tumor therapies.

[Lung protective ventilation - supportive and adjunctive therapies in acute lung injury]

Various supportive and adjunctive therapies to conventional mechanical ventilation have been evaluated in patients with acute lung injury and acute respiratory distress syndrome (e.g. nitric oxide, prone position, surfactant, glucocorticoids). Although some investigations have shown promising improvements in oxygenation and physiological variables, large randomized trials of adjunctive and supportive therapies showed no impact on survival.

Olfactory bulb interneurons releasing NO exhibit the Reelin receptor ApoEr2 and part of those targeted by NO express Reelin

Nitric oxide (NO) and Reelin both modulate neuronal plasticity in developing and mature synaptic networks. We recently showed a loss of neuronal nitric oxide synthase (nNOS) protein in the olfactory bulb of reeler mutants and advanced the hypothesis that the Reelin and NO signalling pathways may influence each other. We now studied the distribution of NO sensitive guanylyl cyclase (NOsGC), Reelin and its receptor Apolipoprotein E2 (ApoEr2) in the olfactory bulb by multiple fluorescence labelling and tested whether nNOS and ApoEr2 colocalize in this area. We also essayed the protein content of NOsGC in the reeler olfactory bulb and tested whether there are any changes in nNOS and NOsGC protein in other reeler brain areas. Olfactory bulb interneurons expressing ApoEr2 and nNOS are only few in the glomerular layer but represent the large majority of granule cell layer interneurons. Conversely, NOsGC interneurons are rare in the granule cell layer and abundant as periglomerular cells. Reelin containing periglomerular cells almost entirely belong to the NOsGC subset. These data further support the hypothesis of a reciprocal signalling between Reelin/NOsGC and ApoEr2/nNOS expressing neurons to affect olfactory bulb activity. We also show that a significant rise in NOsGC content accompanies the decrease of nNOS protein in the reeler olfactory bulb. The same reciprocal changes present in the cortex/striatum and the hippocampus of reeler mice. Thus, the influence that the deficit of extracellular Reelin seems to exert on nNOS and its receptor is not limited to the olfactory bulb but is a general feature of the reeler brain.

[11C]Choline PET/CT for targeted salvage lymph node dissection in patients with biochemical recurrence after primary curative therapy for prostate cancer. Preliminary results of a prospective study

INTRODUCTION: In this prospective study we set out to investigate the diagnostic value of [(11)C]choline-PET/CT in patients with suspected lymph node metastases before salvage lymph node dissection. PATIENTS AND METHODS: 15 consecutive patients with rising PSA underwent [(11)C]choline-PET/CT and consecutive open salvage pelvic/retroperitoneal extended lymph node dissection due to uptake of [(11)C]choline in at least 1 lymph node. Mean age was 62.1 (range 53-73). RESULTS: [(11)C]choline-PET/CT results were compared with the histopathology reports and clinical follow-up (mean 13.7 months, range 6-24). Mean time to progression was 23.6 months (range 4-81). [(11)C]choline uptake was observed in nodes along the external and internal and common iliac arteries and in the paraaortic region. A positive histology was reported in 8/15 patients. Only one patient had a PSA nadir of <0.1 ng/ml after salvage surgery. Another patient had stable disease with a PSA of 0.5 ng/ml. Three patients developed bone metastases during follow-up. CONCLUSIONS: This interim analysis indicates that [(11)C]choline-PET/CT may be a useful technique in detection of lymph node metastases when rising PSA occurs after definite prostate cancer therapy. The presented cohort is limited in size, but there is still strong evidence that the patients benefit from [(11)C]choline-PET/CT and consecutive salvage lymph node dissection is rather small.

Integrin alphavbeta6 is a marker of the progression of biliary and portal liver fibrosis and a novel target for antifibrotic therapies

BACKGROUND/AIMS: The integrin alphavbeta6 promotes proliferation of specialized epithelia and acts as a receptor for the activation of latent TGFbeta1. We studied alphavbeta6 expression in experimental and human liver fibrosis and the potential of its pharmacological inhibition for treatment of hepatic fibrosis. METHODS: alphavbeta6 expression was studied by quantitative PCR and immunohistochemistry in rats with cirrhosis due to bile duct ligation (BDL), administration of thioacetamide (TAA), in Mdr2(Abcb4)(-/-) mice with spontaneous biliary fibrosis, and in livers of patients with chronic hepatitis C (n=79) and end-stage liver disease due to various etiologies (n=18). The effect of a selective alphavbeta6 inhibitor was evaluated in Mdr2(Abcb4)(-/-) mice with ongoing fibrogenesis. RESULTS: Integrin beta6 mRNA increased with fibrosis stage in hepatitis C and was upregulated between 25- and 100-fold in TAA- and BDL-induced cirrhosis, in Mdr2(Abcb4)(-/-) mice and in human end-stage liver disease. alphavbeta6 protein was absent in normal livers and expressed de novo on (activated) bile duct epithelia and transitional hepatocytes. A single dose of the alphavbeta6 inhibitor injected into Mdr2(Abcb4)(-/-) mice significantly induced profibrolytic matrix metalloproteinases (MMP)-8 and -9 after 3 h, with a corresponding increase in extracellular matrix-degrading activities. In parallel profibrogenic transcripts (procollagen alpha1(I), TGFbeta2, and MMP-2) showed a trend of downregulation. CONCLUSIONS: (1) Integrin alphavbeta6 is induced de novo in rodent and human liver fibrosis, where it is expressed on activated bile duct epithelia and (transitional) hepatocytes during fibrosis progression. (2) In vivo a single dose of a small molecule alphavbeta6 inhibitor induced antifibrogenic and profibrolytic genes and activities, suggesting alphavbeta6 is a unique target for treatment of liver fibrosis.

Lipids as targets for novel anti-inflammatory therapies

Lipids serve important functions as membrane constituents and also as energy storing molecules. Besides these functions certain lipid species have now been recognized as signalling molecules that regulate a multitude of cellular responses including cell growth and death, and also inflammatory reactions. Bioactive lipids are generated by hydrolysis from membrane lipids mainly by phospholipases giving rise to fatty acids and lysophospholipids that either directly exert their function or are further converted to active mediators. This review will summarize the present knowledge about bioactive lipids that either promote or attenuate inflammatory reactions. These lipids include polyunsaturated fatty acids (PUFA), eicosanoids including the epoxyeicosatrienoic acids (EET), peroxisome proliferation activating receptor (PPAR) activators, cannabinoids and the sphingolipids ceramide, sphingosine 1-phosphate and sphingosylphosphorylcholine.

Cannabinoid receptor ligands as potential anticancer agents--high hopes for new therapies?

OBJECTIVES: The endocannabinoid system is an endogenous lipid signalling network comprising arachidonic-acid-derived ligands, cannabinoid (CB) receptors, transporters and endocannabinoid degrading enzymes. The CB(1) receptor is predominantly expressed in neurons but is also co-expressed with the CB(2) receptor in peripheral tissues. In recent years, CB receptor ligands, including Delta(9)-tetrahydrocannabinol, have been proposed as potential anticancer agents. KEY FINDINGS: This review critically discusses the pharmacology of CB receptor activation as a novel therapeutic anticancer strategy in terms of ligand selectivity, tissue specificity and potency. Intriguingly, antitumour effects mediated by cannabinoids are not confined to inhibition of cancer cell proliferation; cannabinoids also reduce angiogenesis, cell migration and metastasis, inhibit carcinogenesis and attenuate inflammatory processes. In the last decade several new selective CB(1) and CB(2) receptor agents have been described, but most studies in the area of cancer research have used non-selective CB ligands. Moreover, many of these ligands exert prominent CB receptor-independent pharmacological effects, such as activation of the G-protein-coupled receptor GPR55, peroxisome proliferator-activated receptor gamma and the transient receptor potential vanilloid channels. SUMMARY: The role of the endocannabinoid system in tumourigenesis is still poorly understood and the molecular mechanisms of cannabinoid anticancer action need to be elucidated. The development of CB(2)-selective anticancer agents could be advantageous in light of the unwanted central effects exerted by CB(1) receptor ligands. Probably the most interesting question is whether cannabinoids could be useful in chemoprevention or in combination with established chemotherapeutic agents.

Locally targeted cytoprotection with dextran sulfate attenuates experimental porcine myocardial ischaemia/reperfusion injury

AIMS: Intravascular inflammatory events during ischaemia/reperfusion injury following coronary angioplasty alter and denudate the endothelium of its natural anticoagulant heparan sulfate proteoglycan (HSPG) layer, contributing to myocardial tissue damage. We propose that locally targeted cytoprotection of ischaemic myocardium with the glycosaminoglycan analogue dextran sulfate (DXS, MW 5000) may protect damaged tissue from reperfusion injury by functional restoration of HSPG. METHODS AND RESULTS: In a closed chest porcine model of acute myocardial ischaemia/reperfusion injury (60 min ischaemia, 120 min reperfusion), DXS was administered intracoronarily into the area at risk 5 min prior to reperfusion. Despite similar areas at risk in both groups (39+/-8% and 42+/-9% of left ventricular mass), DXS significantly decreased myocardial infarct size from 61+/-12% of the area at risk for vehicle controls to 39+/-14%. Cardioprotection correlated with reduced cardiac enzyme release creatine kinase (CK-MB, troponin-I). DXS abrogated myocardial complement deposition and substantially decreased vascular expression of pro-coagulant tissue factor in ischaemic myocardium. DXS binding, detected using fluorescein-labelled agent, localized to ischaemically damaged blood vessels/myocardium and correlated with reduced vascular staining of HSPG. CONCLUSION: The significant cardioprotection obtained through targeted cytoprotection of ischaemic tissue prior to reperfusion in this model of acute myocardial infarction suggests a possible role for the local modulation of vascular inflammation by glycosaminoglycan analogues as a novel therapy to reduce reperfusion injury.

Hemodynamic variables and mortality in cardiogenic shock: a retrospective cohort study

INTRODUCTION: Despite the key role of hemodynamic goals, there are few data addressing the question as to which hemodynamic variables are associated with outcome or should be targeted in cardiogenic shock patients. The aim of this study was to investigate the association between hemodynamic variables and cardiogenic shock mortality. METHODS: Medical records and the patient data management system of a multidisciplinary intensive care unit (ICU) were reviewed for patients admitted because of cardiogenic shock. In all patients, the hourly variable time integral of hemodynamic variables during the first 24 hours after ICU admission was calculated. If hemodynamic variables were associated with 28-day mortality, the hourly variable time integral of drops below clinically relevant threshold levels was computed. Regression models and receiver operator characteristic analyses were calculated. All statistical models were adjusted for age, admission year, mean catecholamine doses and the Simplified Acute Physiology Score II (excluding hemodynamic counts) in order to account for the influence of age, changes in therapies during the observation period, the severity of cardiovascular failure and the severity of the underlying disease on 28-day mortality. RESULTS: One-hundred and nineteen patients were included. Cardiac index (CI) (P = 0.01) and cardiac power index (CPI) (P = 0.03) were the only hemodynamic variables separately associated with mortality. The hourly time integral of CI drops <3, 2.75 (both P = 0.02) and 2.5 (P = 0.03) L/min/m2 was associated with death but not that of CI drops <2 L/min/m2 or lower thresholds (all P > 0.05). The hourly time integral of CPI drops <0.5-0.8 W/m2 (all P = 0.04) was associated with 28-day mortality but not that of CPI drops <0.4 W/m2 or lower thresholds (all P > 0.05). CONCLUSIONS: During the first 24 hours after intensive care unit admission, CI and CPI are the most important hemodynamic variables separately associated with 28-day mortality in patients with cardiogenic shock. A CI of 3 L/min/m2 and a CPI of 0.8 W/m2 were most predictive of 28-day mortality. Since our results must be considered hypothesis-generating, randomized controlled trials are required to evaluate whether targeting these levels as early resuscitation endpoints can improve mortality in cardiogenic shock.