Utility of a simplified ultrasound assessment to assess interstitial pulmonary fibrosis in connective tissue disorders - preliminary results

Introduction  

Interstitial pulmonary fibrosis (IPF) is a frequent manifestation in patients with connective tissue disorders (CTD). Recently the ultrasound (US) criterion validity for its assessment has been proposed; however, the US scoring systems adopted include the study of several lung intercostal spaces (LIS), which could be time-consuming in daily clinical practice. The aim of this study was to investigate the utility of a simplified US B-lines scoring system compared with both the US comprehensive assessment and the high-resolution computed tomography (HRCT) findings of IPF in CTD patients.     

Local tissue interactions across the dorsal midline of the forebrain establish CNS laterality

The mechanisms that establish behavioral, cognitive, and neuroanatomical asymmetries are poorly understood. In this study, we analyze the events that regulate development of asymmetric nuclei in the dorsal forebrain. The unilateral parapineal organ has a bilateral origin, and some parapineal precursors migrate across the midline to form this left-sided nucleus. The parapineal subsequently innervates the left habenula, which derives from ventral epithalamic cells adjacent to the parapineal precursors. Ablation of cells in the left ventral epithalamus can reverse laterality in wild-type embryos and impose the direction of CNS asymmetry in embryos in which laterality is usually randomized. Unilateral modulation of Nodal activity by Lefty1 can also impose the direction of CNS laterality in embryos with bilateral expression of Nodal pathway genes. From these data, we propose that laterality is determined by a competitive interaction between the left and right epithalamus and that Nodal signaling biases the outcome of this competition.     

Abnormal lymphatic vessel development in neuropilin 2 mutant mice

Neuropilin 2 is a receptor for class III semaphorins and for certain members of the vascular endothelial growth factor family. Targeted inactivation of the neuropilin 2 gene (Nrp2) has previously shown its role in neural development. We report that neuropilin 2 expression in the vascular system is restricted to veins and lymphatic vessels. Homozygous Nrp2 mutants show absence or severe reduction of small lymphatic vessels and capillaries during development. This correlated with a reduction of DNA synthesis in the lymphatic endothelial cells of the mutants. Arteries, veins and larger, collecting lymphatic vessels developed normally, suggesting that neuropilin 2 is selectively required for the formation of small lymphatic vessels and capillaries.     

Cloning of bovine LYST gene and identification of a missense mutation associated with Chediak-Higashi syndrome of cattle

An inheritable bleeding disorder with light coat color caused by an autosomal recessive gene has been reported in a population of Japanese black cattle. The disease has been diagnosed as Chediak-Higashi Syndrome (CHS) of cattle which correspond to a human inheritable disorder caused by mutation in LYST gene. To characterize the molecular lesion causing CHS in cattle, cDNAs encoding bovine LYST were isolated from a bovine brain cDNA library. The nucleotide and deduced amino acid sequences of bovine LYST had 89.6 and 90.2% identity with those of the human LYST gene, respectively. In order to identify the mutation within the LYST gene causing CHS in cattle, cDNA fragments of the LYST gene were amplified from an affected animal by RT-PCR and their nucleotide sequences were completely determined. Notably, a nucleotide substitution of A to G transition, resulting in an amino acid substitution of histidine to arginine (H2015R) was identified in the affected animal. The presence of the substitution was completely corresponding with the occurrence of the CHS phenotype among 105 members of pedigrees of the Japanese black cattle and no cattle of other populations had this substitution. These findings strongly suggested that H2015R is the causative mutation in CHS of Japanese black cattle. Received: 25 May 1999 / Accepted: 26 July 1999     

Fluorone dyes have binding sites on both cytoplasmic and extracellular domains of Na,K-ATPase

Combination of fluorescence techniques and molecular docking was used to monitor interaction of Na,K-ATPase and its large cytoplasmic loop connecting fourth and fifth transmembrane helices (C45) with fluorone dyes (i.e. eosin Y, 5(6)-carboxyeosin, rose bengal, fluorescein, and erythrosine B). Our data suggested that there are at least two binding sites for all used fluorone dyes, except of 5(6)-carboxyeosin. The first binding site is located on C45 loop, and it is sensitive to the presence of nucleotide. The other site is located on the extracellular part of the enzyme, and it is sensitive to the presence of Na⁺ or K⁺ ions. The molecular docking revealed that in the open conformation of C45 loop (which is obtained in the presence of ATP) all used fluorone dyes occupy position directly inside the ATP-binding pocket, while in the closed conformation (i.e. in the absence of any ligand) they are located only near the ATP-binding site depending on their different sizes. On the extracellular part of the protein, the molecular docking predicts two possible binding sites with similar binding energy near Asp897(α) or Gln69(β). The former was identified as a part of interaction site between α- and β-subunits, the latter is in contact with conserved FXYD sequence of the γ-subunit. Our findings provide structural explanation for numerous older studies, which were performed with fluorone dyes before the high-resolution structures were known. Further, fluorone dyes seem to be good probes for monitoring of intersubunit interactions influenced by Na⁺ and K⁺ binding.     

A meta-analysis of the life cycle greenhouse gas balances of microalgae biodiesel

Purpose

Microalgae biodiesel has attracted considerable attention as a potential substitute for fossil fuels and biodiesel from food crops. Nevertheless, its reported climate impacts in the scientific literature vary significantly. This article describes and synthesizes the range of results found in the life cycle assessment (LCA) literature regarding microalgae biodiesel studies to investigate whether particular parameters, e.g. technologies, were associated with higher or lower greenhouse gas (GHG) emissions so that a best practice can be inferred from currently available LCA data and thereby recommended.

Methods

A systematic literature review and meta-regression analysis (MRA) of 36 LCA studies that report on the GHG emissions of microalgae biodiesel was conducted. An assessment of key aspects, including modelling choices and technologies, was performed. Furthermore, MRA models were formulated considering several variables of interest describing both technical and modelling choices to identify the main causes for the variability in GHG emissions per MJ of biodiesel. Variables chosen include: microalgae species; culture medium; cultivation system; source of CO₂; extraction technology; conversion technology; system boundary; geographical scope; inclusion or exclusion of capital goods; and how multifunctionality was handled.

Results and discussion

The reviewed studies altogether reported 308 results ranging from ?0.7 to 3.8 kg CO₂ eq. MJ^?1_biodiesel, portraying 19 different system configurations. Despite the comprehensive range of variables assessed, the models generated could not plausibly explain that the variability in GHG emissions depends either on the technologies considered or on the methodological choices adopted. However, the following relationships could be observed: location in Europe and high oil productivity were associated with lower emissions, whilst dry extraction should be avoided for leading to higher GHG emissions, on average.

Conclusions

There is a large degree of variability within the technologies considered, as well as the methodological choices adopted, so that no robust conclusions could be drawn from the MRA. Notwithstanding, average GHG emissions reported were more than twice as high as fossil diesel and, while there are some studies showing large benefits, none of the various algae technologies performed consistently better than fossil diesel, questioning the climate-mitigation potential of microalgae biodiesel.

     

One drug for two targets: Biological evaluation of antiretroviral agents endowed with antiproliferative activity

AIDS-related cancer diseases are malignancies with low incidence on healthy people that affect mostly subjects already immunocompromised. The connection between HIV/AIDS and these cancers has not been established yet, but a weakened immune system is certainly the main cause. We envisaged the possibility to screen a small library of compounds synthesized in our laboratory against opportunistic tumors mainly due to HIV infection like Burkitt’s Lymphoma. From cellular assays and gene expression analysis we identified two promising compounds. These derivatives have the dual action required inhibiting HIV replication in human TZM-bl cells infected with HIV-1 NL4.3 and showing cytotoxic activity on human colon HT-29 and breast adenocarcinoma MCF-7 cells. In addition, preclinical in vitro adsorption, distribution, metabolism, and excretion studies highlighted a satisfactory pharmacokinetic profile.     

Establishment of a hepatitis C virus subgenomic replicon derived from human hepatocytes infected in vitro

The hepatitis C virus (HCV) replicon system is a potent tool for understanding the mechanisms of HCV replication and proliferation, and for the development of treatments for patients with HCV. Recently, we established an HCV subgenomic replicon (50-1) using HCV genome RNA obtained from the cultured human T cell line MT-2C infected with HCV (isolate 1B-1) in vitro. In order to further obtain other HCV replicons without difficulty, we generated a replicon RNA library derived from human non-neoplastic hepatocytes infected with HCV (isolate 1B-2) in vitro. Upon transfection of the generated RNA library to "cured cells," from which the 50-1 subgenomic replicon was eliminated by prolonged treatment with interferon-alpha, we successfully established a new HCV subgenomic replicon, 1B-2R1. We characterized 1B-2R1 replicon in terms of efficiency of replication, HCV sequence, and sensitivity to interferons. The results revealed that the replication level of the 1B-2R1 replicon was comparable to that of the 50-1 replicon. We also found that the 1B-2R1 replicon possessed an HCV sequence distinct from those of other replicons established to date, and that the 1B-2R1 replicon was sensitive to interferon-alpha, interferon-beta, and interferon-gamma. Taken together, present results indicate that the replicon RNA library generated using an in vitro HCV infection system is useful for the establishment of an HCV subgenomic replicon.     

Association of simian virus 40 vp1 with 70-kilodalton heat shock proteins and viral tumor antigens

Proper folding of newly synthesized viral proteins in the cytoplasm is a prerequisite for the formation of infectious virions. The major capsid protein Vp1 of simian virus 40 forms a series of disulfide-linked intermediates during folding and capsid formation. In addition, we report here that Vp1 is associated with cellular chaperones (HSP70) and a cochaperone (Hsp40) which can be coimmunoprecipitated with Vp1. Studies in vitro demonstrated the ATP-dependent interaction of Vp1 and cellular chaperones. Interestingly, viral cochaperones LT and ST were essential for stable interaction of HSP70 with the core Vp1 pentamer Vp1 (22-303). LT and ST also coimmunoprecipitated with Vp1 in vivo. In addition to these identified (co)chaperones, stable, covalently modified forms of Vp1 were identified for a folding-defective double mutant, C49A-C87A, and may represent a “trapped” assembly intermediate. By a truncation of the carboxyl arm of Vp1 to prevent the Vp1 folding from proceeding beyond pentamers, we detected several apparently modified Vp1 species, some of which were absent in cells transfected with the folding-defective mutant DNA. These results suggest that transient covalent interactions with known or unknown cellular and viral proteins are important in the assembly process.