The pharmacokinetics and safety of porfimer after repeated administration 30-45 days apart to patients undergoing photodynamic therapy

Porfimer is an intravenous (i.v.) injectable photosensitizing agent used in the photodynamic treatment of tumours and of high-grade dysplasia in Barrett's oesophagus.

Level of G protein-coupled receptor kinase-2 determines myocardial ischemia/reperfusion injury via pro- and anti-apoptotic mechanisms

Activation of prosurvival kinases and subsequent nitric oxide (NO) production by certain G protein-coupled receptors (GPCRs) protects myocardium in ischemia/reperfusion injury (I/R) models. GPCR signaling pathways are regulated by GPCR kinases (GRKs), and GRK2 has been shown to be a critical molecule in normal and pathological cardiac function.

Vulnerability to risk among small farmers in Tajikistan: results of a 2011 survey

Tajikistan is judged to be highly vulnerable to risk, including food insecurity risks and climate change risks. By some vulnerability measures it is the most vulnerable among all 28 countries in the World Bank’s Europe and Central Asia Region – ECA (World Bank 2009). The rural population, with its relatively high incidence of poverty, is particularly vulnerable. The Pilot Program for Climate Resilience (PPCR) in Tajikistan (2011) provided an opportunity to conduct a farm-level survey with the objective of assessing various dimensions of rural population’s vulnerability to risk and their perception of constraints to farming operations and livelihoods. The survey should be accordingly referred to as the 2011 PPCR survey. The rural population in Tajikistan is highly agrarian, with about 50% of family income deriving from agriculture (see Figure 4.1; also LSMS 2007 – own calculations). Tajikistan’s agriculture basically consists of two groups of producers: small household plots – the successors of Soviet “private agriculture” – and dehkan (or “peasant”) farms – new family farming structures that began to be created under relevant legislation passed after 1992 (Lerman and Sedik, 2008). The household plots manage 20% of arable land and produce 65% of gross agricultural output (GAO). Dehkan farms manage 65% of arable land and produce close to 30% of GAO. The remaining 15% of arable land is held in agricultural enterprises – the rapidly shrinking sector of corporate farms that succeeded the Soviet kolkhozes and sovkhozes and today produces less than 10% of GAO (TajStat 2011) The survey conducted in May 2011 focused on dehkan farms, as budgetary constraints precluded the inclusion of household plots. A total of 142 dehkan farms were surveyed in face-to-face interviews. They were sampled from 17 districts across all four regions – Sughd, Khatlon, RRP, and GBAO. The districts were selected so as to represent different agro-climatic zones, different vulnerability zones (based on the World Bank (2011) vulnerability assessment), and different food-insecurity zones (based on WFP/IPC assessments). Within each district, 3-4 jamoats were chosen at random and 2-3 farms were selected in each jamoat from lists provided by jamoat administration so as to maximize the variability by farm characteristics. The sample design by region/district is presented in Table A, which also shows the agro-climatic zone and the food security phase for each district. The sample districts are superimposed on a map of food security phases based on IPC April 2011.

Sleep disturbance impairs stroke recovery in the rat

There is a lack of experimental evidence to support the hypothesis that sleep may modulate stroke outcome as suggested by clinical observations. We have previously shown that sleep disturbance (SDis) over 3 days aggravates brain damage in a rat model of focal cerebral ischemia. The aim of this study is to further investigate effects of SDis on long-term stroke recovery and neuroplasticity as assessed by axonal sprouting, neurogenesis, and angiogenesis.

Cardiac G-protein-coupled receptor kinase 2 ablation induces a novel Ca2+ handling phenotype resistant to adverse alterations and remodeling after myocardial infarction

G-protein-coupled receptor kinase 2 (GRK2) is a primary regulator of β-adrenergic signaling in the heart. G-protein-coupled receptor kinase 2 ablation impedes heart failure development, but elucidation of the cellular mechanisms has not been achieved, and such elucidation is the aim of this study.

Lead discovery, chemistry optimization, and biological evaluation studies of novel biamide derivatives as CB2 receptor inverse agonists and osteoclast inhibitors

N,N'-((4-(Dimethylamino)phenyl)methylene)bis(2-phenylacetamide) was discovered by using 3D pharmacophore database searches and was biologically confirmed as a new class of CB(2) inverse agonists. Subsequently, 52 derivatives were designed and synthesized through lead chemistry optimization by modifying the rings A-C and the core structure in further SAR studies. Five compounds were developed and also confirmed as CB(2) inverse agonists with the highest CB(2) binding affinity (CB(2)K(i) of 22-85 nM, EC(50) of 4-28 nM) and best selectivity (CB(1)/CB(2) of 235- to 909-fold). Furthermore, osteoclastogenesis bioassay indicated that PAM compounds showed great inhibition of osteoclast formation. Especially, compound 26 showed 72% inhibition activity even at the low concentration of 0.1 μM. The cytotoxicity assay suggested that the inhibition of PAM compounds on osteoclastogenesis did not result from its cytotoxicity. Therefore, these PAM derivatives could be used as potential leads for the development of a new type of antiosteoporosis agent.

Sleep deprivation before stroke is neuroprotective: A pre-ischemic conditioning related to sleep rebound

BACKGROUND AND AIM: We have previously shown in a rat model of focal cerebral ischemia that sleep deprivation after stroke onset aggravates brain damage. Others reported that sleep deprivation prior to stroke is neuroprotective. The main aim of this study was to test the hypothesis that the neuroprotection may be related to an increase in sleep (sleep rebound) during the acute phase of stroke. METHODS: Male Sprague Dawley rats (n=36) were subjected to continuous polygraphic recordings for baseline, total sleep deprivation (TSD), and 24h after ischemia. TSD for 6h was performed by gentle handling and immediately followed by ischemia. Focal cerebral ischemia was induced by permanent occlusion of distal branches of the middle cerebral artery. Control experiments included ischemia without SD (nSD) and sham surgery with TSD (n=6/group). RESULTS: Shortly after stroke, the amount of slow wave sleep (SWS) and paradoxical sleep (PS) increased significantly (p<0.05) in the TSD/ischemia, resulting in an increase in the total sleep time by 30% compared to baseline, or by 20% compared with the nSD/ischemia group. The infarct volume decreased significantly by 50% in the TSD/ischemia compared to nSD group (p<0.02). Removal of sleep rebound by allowing TSD-rats sleep for 24h before ischemia eliminated the reduction in the infarct size. CONCLUSION PRESTROKE: Sleep deprivation results in sleep rebound and reduces brain damage. Sleep rebound may be causally related to the neuroprotection.

Guidelines for the use and interpretation of assays for monitoring autophagy in higher eukaryotes

Research in autophagy continues to accelerate,(1) and as a result many new scientists are entering the field. Accordingly, it is important to establish a standard set of criteria for monitoring macroautophagy in different organisms. Recent reviews have described the range of assays that have been used for this purpose.(2,3) There are many useful and convenient methods that can be used to monitor macroautophagy in yeast, but relatively few in other model systems, and there is much confusion regarding acceptable methods to measure macroautophagy in higher eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers of autophagosomes versus those that measure flux through the autophagy pathway; thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from fully functional autophagy that includes delivery to, and degradation within, lysosomes (in most higher eukaryotes) or the vacuole (in plants and fungi). Here, we present a set of guidelines for the selection and interpretation of the methods that can be used by investigators who are attempting to examine macroautophagy and related processes, as well as by reviewers who need to provide realistic and reasonable critiques of papers that investigate these processes. This set of guidelines is not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to verify an autophagic response.

Synthesis and in vitro characterization of radioiodinatable benzodiazepines selective for type 1 and type 2 cholecystokinin receptors

Radiolabeled antagonists of specific peptide receptors identify a higher number of receptor binding sites than agonists and may thus be preferable for in vivo tumor targeting. In this study, two novel radioiodinated 1,4-benzodiazepines, (S)-1-(3-iodophenyl)-3-(1-methyl-2-oxo-5-phenyl-2,3-dihydro-1H-benzo[e][1,4]diazepin-3-yl)urea (9) and (R)-1-(3-iodophenyl)-3-(1-methyl-2-oxo-5-phenyl-2,3-dihydro-1H-benzo[e][1,4]diazepin-3-yl)urea (7), were developed. They were characterized in vitro as high affinity selective antagonists at cholecystokinin types 1 and 2 (CCK(1) and CCK(2)) receptors using receptor binding, calcium mobilization, and internalization studies. Their binding to human tumor tissues was assessed with in vitro receptor autoradiography and compared with an established peptidic CCK agonist radioligand. The (125)I-labeled CCK(1) receptor-selective compound 9 often revealed a substantially higher amount of CCK(1) receptor binding sites in tumors than the agonist (125)I-CCK. Conversely, the radioiodinated CCK(2) receptor-selective compound 7 showed generally weaker tumor binding than (125)I-CCK. In conclusion, compound 9 is an excellent radioiodinated nonpeptidic antagonist ligand for direct and selective labeling of CCK(1) receptors in vitro. Moreover, it represents a suitable candidate to test antagonist binding to CCK(1) receptor-expressing tumors in vivo.

Identification of small Sca-1(+), Lin(-), CD45(-) multipotential cells in the neonatal murine retina

OBJECTIVE: Bone marrow contains a subset of stem cells that give rise to nonhematopoietic lineages. These nonhematopoietic stem cells appear heterogeneous and contain cells committed to mesenchymal and endothelial lineages, as well as more primitive multipotential cells resembling progenitors of germ cells and very small embryonic/epiblast-like stem cells (VSELs). Nonhematopoietic stem cells can be mobilized from the bone marrow in response to tissue injury, and cells with similar properties have been found in cord blood and normal adult organs. However, the relationship between bone marrow cells and these adult organ stem cells is still unclear. The differentiation potential of some adult stem cells is organ-restricted, but other populations appear to retain multipotential capacity. MATERIALS AND METHODS: A population of small Sca-1(+), lineage-negative (Lin(-)), CD45(-) cells resembling VSELs were isolated from neonatal mouse retina by cell sorting. Differentiation of the cells in culture was achieved by exposure to embryonic stem cell differentiation protocols. RESULTS: VSEL-like cells comprise 1.5% of the neonatal mouse retina. They remain quiescent during retinal differentiation, and thus they do not contribute to normal retinal development. However, they display eye cell differentiation potential in culture and they are also multipotential and can give rise to cells representative of all three embryonic layers. CONCLUSIONS: The neonatal retina is an abundant postnatal source of multipotential VSEL-like cells that can differentiate in culture into a variety of lineages.

Myocardial Ablation of G Protein-Coupled Receptor Kinase 2 (GRK2) Decreases Ischemia/Reperfusion Injury through an Anti-Intrinsic Apoptotic Pathway

Studies from our lab have shown that decreasing myocardial G protein-coupled receptor kinase 2 (GRK2) activity and expression can prevent heart failure progression after myocardial infarction. Since GRK2 appears to also act as a pro-death kinase in myocytes, we investigated the effect of cardiomyocyte-specific GRK2 ablation on the acute response to cardiac ischemia/reperfusion (I/R) injury. To do this we utilized two independent lines of GRK2 knockout (KO) mice where the GRK2 gene was deleted in only cardiomyocytes either constitutively at birth or in an inducible manner that occurred in adult mice prior to I/R. These GRK2 KO mice and appropriate control mice were subjected to a sham procedure or 30 min of myocardial ischemia via coronary artery ligation followed by 24 hrs reperfusion. Echocardiography and hemodynamic measurements showed significantly improved post-I/R cardiac function in both GRK2 KO lines, which correlated with smaller infarct sizes in GRK2 KO mice compared to controls. Moreover, there was significantly less TUNEL positive myocytes, less caspase-3, and -9 but not caspase-8 activities in GRK2 KO mice compared to control mice after I/R injury. Of note, we found that lowering cardiac GRK2 expression was associated with significantly lower cytosolic cytochrome C levels in both lines of GRK2 KO mice after I/R compared to corresponding control animals. Mechanistically, the anti-apoptotic effects of lowering GRK2 expression were accompanied by increased levels of Bcl-2, Bcl-xl, and increased activation of Akt after I/R injury. These findings were reproduced in vitro in cultured cardiomyocytes and GRK2 mRNA silencing. Therefore, lowering GRK2 expression in cardiomyocytes limits I/R-induced injury and improves post-ischemia recovery by decreasing myocyte apoptosis at least partially via Akt/Bcl-2 mediated mitochondrial protection and implicates mitochondrial-dependent actions, solidifying GRK2 as a pro-death kinase in the heart.

Inhibition of Fas-associated death domain-containing protein (FADD) protects against myocardial ischemia/reperfusion injury in a heart failure mouse model

AIM As technological interventions treating acute myocardial infarction (MI) improve, post-ischemic heart failure increasingly threatens patient health. The aim of the current study was to test whether FADD could be a potential target of gene therapy in the treatment of heart failure. METHODS Cardiomyocyte-specific FADD knockout mice along with non-transgenic littermates (NLC) were subjected to 30 minutes myocardial ischemia followed by 7 days of reperfusion or 6 weeks of permanent myocardial ischemia via the ligation of left main descending coronary artery. Cardiac function were evaluated by echocardiography and left ventricular (LV) catheterization and cardiomyocyte death was measured by Evans blue-TTC staining, TUNEL staining, and caspase-3, -8, and -9 activities. In vitro, H9C2 cells transfected with ether scramble siRNA or FADD siRNA were stressed with chelerythrin for 30 min and cleaved caspase-3 was assessed. RESULTS FADD expression was significantly decreased in FADD knockout mice compared to NLC. Ischemia/reperfusion (I/R) upregulated FADD expression in NLC mice, but not in FADD knockout mice at the early time. FADD deletion significantly attenuated I/R-induced cardiac dysfunction, decreased myocardial necrosis, and inhibited cardiomyocyte apoptosis. Furthermore, in 6 weeks long term permanent ischemia model, FADD deletion significantly reduced the infarct size (from 41.20 ± 3.90% in NLC to 26.83 ± 4.17% in FADD deletion), attenuated myocardial remodeling, improved cardiac function and improved survival. In vitro, FADD knockdown significantly reduced chelerythrin-induced the level of cleaved caspase-3. CONCLUSION Taken together, our results suggest FADD plays a critical role in post-ischemic heart failure. Inhibition of FADD retards heart failure progression. Our data supports the further investigation of FADD as a potential target for genetic manipulation in the treatment of heart failure.

Glycoprotein biosynthesis in a eukaryote lacking the membrane protein Rft1

Mature dolichol-linked oligosaccharides (mDLOs) needed for eukaryotic protein N-glycosylation are synthesized by a multistep pathway in which the biosynthetic lipid intermediate Man5GlcNAc2-PP-dolichol (M5-DLO) flips from the cytoplasmic to the luminal face of the endoplasmic reticulum. The endoplasmic reticulum membrane protein Rft1 is intimately involved in mDLO biosynthesis. Yeast genetic analyses implicated Rft1 as the M5-DLO flippase, but because biochemical tests challenged this assignment, the function of Rft1 remains obscure. To understand the role of Rft1, we sought to analyze mDLO biosynthesis in vivo in the complete absence of the protein. Rft1 is essential for yeast viability, and no Rft1-null organisms are currently available. Here, we exploited Trypanosoma brucei (Tb), an early diverging eukaryote whose Rft1 homologue functions in yeast. We report that TbRft1-null procyclic trypanosomes grow nearly normally. They have normal steady-state levels of mDLO and significant N-glycosylation, indicating robust M5-DLO flippase activity. Remarkably, the mutant cells have 30-100-fold greater steady-state levels of M5-DLO than wild-type cells. All N-glycans in the TbRft1-null cells originate from mDLO indicating that the M5-DLO excess is not available for glycosylation. These results suggest that rather than facilitating M5-DLO flipping, Rft1 facilitates conversion of M5-DLO to mDLO by another mechanism, possibly by acting as an M5-DLO chaperone.

The role of subpolar deep water formation and Nordic Seas overflows in simulated multidecadal variability of the Atlantic meridional overturning circulation

We investigate the respective role of variations in subpolar deep water formation and Nordic Seas overflows for the decadal to multidecadal variability of the Atlantic meridional overturning circulation (AMOC). This is partly done by analysing long (order of 1000 years) control simulations with five coupled climate models. For all models, the maximum influence of variations in subpolar deep water formation is found at about 45° N, while the maximum influence of variations in Nordic Seas overflows is rather found at 55 to 60° N. Regarding the two overflow branches, the influence of variations in the Denmark Strait overflow is, for all models, substantially larger than that of variations in the overflow across the Iceland–Scotland Ridge. The latter might, however, be underestimated, as the models in general do not realistically simulate the flow path of the Iceland–Scotland overflow water south of the Iceland–Scotland Ridge. The influence of variations in subpolar deep water formation is, on multimodel average, larger than that of variations in the Denmark Strait overflow. This is true both at 45° N, where the maximum standard deviation of decadal to multidecadal AMOC variability is located for all but one model, and at the more classical latitude of 30° N. At 30° N, variations in subpolar deep water formation and Denmark Strait overflow explain, on multimodel average, about half and one-third respectively of the decadal to multidecadal AMOC variance. Apart from analysing multimodel control simulations, we have performed sensitivity experiments with one of the models, in which we suppress the variability of either subpolar deep water formation or Nordic Seas overflows. The sensitivity experiments indicate that variations in subpolar deep water formation and Nordic Seas overflows are not completely independent. We further conclude from these experiments that the decadal to multidecadal AMOC variability north of about 50° N is mainly related to variations in Nordic Seas overflows. At 45° N and south of this latitude, variations in both subpolar deep water formation and Nordic Seas overflows contribute to the AMOC variability, with neither of the processes being very dominant compared to the other.