Involvement of caspase-dependent activation of cytosolic phospholipase A2 in tumor necrosis factor-induced apoptosis

Tumor necrosis factor (TNF)-induced apoptosis is mediated by caspases, which are cysteine proteases related to interleukin 1β-converting enzyme. We report here that TNF-induced activation of caspases results in the cleavage and activation of cytosolic phospholipase A2 (cPLA2) and that activated cPLA2 contributes to apoptosis. Inhibition of caspases by expression of a cowpox virus-derived inhibitor, CrmA, or by a specific tetrapeptide inhibitor of CPP32/caspase-3, acetyl-Asp-Glu-Val-Asp-aldehyde (Ac-DEVD-CHO), inhibited TNF-induced activation of cPLA2 and apoptosis. TNF-induced activation of cPLA2 was accompanied by a cleavage of the 100-kDa cPLA2 to a 70-kDa proteolytic fragment. This cleavage was inhibited by Ac-DEVD-CHO in a similar manner as that of poly(ADP)ribose polymerase, a known substrate of CPP32/caspase-3. Interestingly, specific inhibition of cPLA2 enzyme activity by arachidonyl trifluoromethylketone (AACOCF3) partially inhibited TNF-induced apoptosis without inhibition of caspase activity. Thus, our results suggest a novel caspase-dependent activation pathway for cPLA2 during apoptosis and identify cPLA2 as a mediator of TNF-induced cell death acting downstream of caspases.

Folding of Active β-Lactamase in the Yeast Cytoplasm before Translocation into the Endoplasmic Reticulum

Polypeptides targeted to the yeast endoplasmic reticulum (ER) posttranslationally are thought to be kept in the cytoplasm in an unfolded state by Hsp70 chaperones before translocation. We show here that Escherichia coli β-lactamase associated with Hsp70, but adopted a native-like conformation before translocation in living Saccharomyces cerevisiae cells. β-Lactamase is a globular trypsin-resistant molecule in authentic form. For these studies, it was linked to the C terminus of a yeast polypeptide Hsp150Δ, which conferred posttranslational translocation and provided sites for O-glycosylation. We devised conditions to retard translocation of Hsp150Δ-β-lactamase. This enabled us to show by protease protection assays that an unglycosylated precursor was associated with the cytoplasmic surface of isolated microsomes, whereas a glycosylated form resided inside the vesicles. Both proteins were trypsin resistant and had similar β-lactamase activity and Km values for nitrocefin. The enzymatically active cytoplasmic intermediate could be chased into the ER, followed by secretion of the activity to the medium. Productive folding in the cytoplasm occurred in the absence of disulfide formation, whereas in the ER lumen, proper folding required oxidation of the sulfhydryls. This suggests that the polypeptide was refolded in the ER and consequently, at least partially unfolded for translocation.

The Cytoplasmic Chaperone Hsp104 Is Required for Conformational Repair of Heat-denatured Proteins in the Yeast Endoplasmic Reticulum

Severe heat stress causes protein denaturation in various cellular compartments. If Saccharomyces cerevisiae cells grown at 24°C are preconditioned at 37°C, proteins denatured by subsequent exposure to 48–50°C can be renatured when the cells are allowed to recover at 24°C. Conformational repair of vital proteins is essential for survival, because gene expression is transiently blocked after the thermal insult. Refolding of cytoplasmic proteins requires the Hsp104 chaperone, and refolding of lumenal endoplasmic reticulum (ER) proteins requires the Hsp70 homologue Lhs1p. We show here that conformational repair of heat-damaged glycoproteins in the ER of living yeast cells required functional Hsp104. A heterologous enzyme and a number of natural yeast proteins, previously translocated and folded in the ER and thereafter denatured by severe heat stress, failed to be refolded to active and secretion-competent structures in the absence of Hsp104 or when an ATP-binding site of Hsp104 was mutated. During recovery at 24°C, the misfolded proteins persisted in the ER, although the secretory apparatus was fully functional. Hsp104 appears to control conformational repair of heat-damaged proteins even beyond the ER membrane.

The adaptor protein Crk connects multiple cellular stimuli to the JNK signaling pathway

c-Jun N-terminal kinases (JNKs) are potently activated by a number of cellular stimuli. Small GTPases, in particular Rac, are responsible for initiating the activation of the JNK pathways. So far, the signals leading from extracellular stimuli to the activation of Rac have remained elusive. Recent studies have demonstrated that the Src homology 2 (SH2)- and Src homology 3 (SH3)-containing adaptor protein Crk is capable of activating JNK when ectopically expressed. We found here that transient expression of Crk induces JNK activation, and this activation was dependent on both the SH2- and SH3-domains of Crk. Expression of p130Cas (Cas), a major binding protein for the Crk SH2-domain, also induced JNK activation, which was blocked by the SH2-mutant of Crk. JNK activation by Cas and Crk was effectively blocked by a dominant-negative form of Rac, suggesting for a linear pathway from the Cas-Crk-complex to the Rac-JNK activation. Many of the stimuli that activate the Rac-JNK pathway enhance engagement of the Crk SH2-domain. JNK activation by these stimuli, such as epidermal growth factor, integrin ligand binding and v-Src, was efficiently blocked by dominant-negative mutants of Crk. A dominant-negative form of Cas in turn blocked the integrin-, but not epidermal growth factor - nor v-Src-mediated JNK activation. Together, these results demonstrate an important role for Crk in connecting multiple cellular stimuli to the Rac-JNK pathway, and a role for the Cas-Crk complex in integrin-mediated JNK activation.

Percutaneous removal of a knotted pulmonary artery catheter using a tracheostomy dilator

This case report describes removal of a knotted, subclavian, pulmonary artery catheter using a tracheostomy dilator. With this simple method an invasive procedure might be averted.

Quaternary ammonium-functionalized silica sorbents for the solid-phase extraction of aromatic amines under normal phase conditions

Quaternary ammonium-functionalized silica materials were synthesized and applied for solid-phase extraction (SPE) of aromatic amines, which are classified as priority pollutants by US Environmental Protection Agency. Hexamethylenetetramine used for silica surface modification for the first time was employed as SPE sorbent under normal phase conditions. Hexaminium-functionalized silica demonstrated excellent extraction efficiencies for o-toluidine, 4-ethylaniline and quinoline (recoveries 101–107%), while for N,N-dimethylaniline and N-isopropylaniline recoveries were from low to moderate (14–46%). In addition, the suitability of 1-alkyl-3-(propyl-3-sulfonate) imidazolium-functionalized silica as SPE sorbent was tested under normal phase conditions. The recoveries achieved for the five aromatic amines ranged from 89 to 99%. The stability of the sorbent was evaluated during and after 150 extractions. Coefficients of variation between 4.5 and 10.2% proved a high stability of the synthesized sorbent. Elution was carried out using acetonitrile in the case of hexaminium-functionalized silica and water for 1-alkyl-3-(propyl-3-sulfonate) imidazolium-functionalized silica sorbent. After the extraction the analytes were separated and detected by liquid chromatography ultraviolet detection (LC-UV). The retention mechanism of the materials was primarily based on polar hydrogen bonding and π–π interactions. Comparison made with activated silica proved the quaternary ammonium-functionalized materials to offer different selectivity and better extraction efficiencies for aromatic amines. Finally, 1-alkyl-3-(propyl-3-sulfonate) imidazolium-functionalized silica sorbent was successfully tested for the extraction of wastewater and soil samples.

Zooplankton biomass from METEOR cruise M87/1 in April 2012

Zooplankton samples were taken in five depth strata using a Multinet type Midi, with 50 µm nets. The samples were taken during the second leg only, three times at station 1, two times at station 2 and once at station 3. Zooplankton were identified to species / genus and life-stage, and at least 300 individuals were counted per sample. 10 individuals of each stage / species were measured and the numbers of eggs counted.

Reducing suffering in animal models and procedures involving seizures, convulsions and epilepsy

This report is based on discussions and submissions from an expert working group consisting of veterinarians, animal care staff and scientists with expert knowledge relevant to the field and aims to facilitate the implementation of the Three Rs (replacement, reduction and refinement) in the use of animal models or procedures involving seizures, convulsions and epilepsy. Each of these conditions will be considered, the specific welfare issues discussed, and practical measures to reduce animal use and suffering suggested. The emphasis is on refinement since this has the greatest potential for immediate implementation, and some general issues for refinement are summarised to help achieve this, with more detail provided on a range of specific refinements.

Reducing suffering in experimental autoimmune encephalomyelitis (EAE)

This report is based on discussions and submissions from an expert working group consisting of veterinarians, animal care staff and scientists with expert knowledge relevant to the field. It aims to facilitate the implementation of the Three Rs (replacement, reduction and refinement) in the use of animal models or procedures involving experimental autoimmune encephalomyelitis (EAE), an experimental model used in multiple sclerosis research. The emphasis is on refinement since this has the greatest potential for immediate implementation. Specific welfare issues are identified and discussed, and practical measures are proposed to reduce animal use and suffering. Some general issues for refinement are summarised to help achieve this, with more detail provided on a range of specific measures to reduce suffering. © 2013 Elsevier Inc.

Technology Vendors: Lodging Managers View Support They Receive

The authors report on a comparative study of regional differences in the perceptions of lodging managers in the United States, Canada, and the United Kingdom on the support they receive from their technology vendors, and the technology systems they are using. Besides a comparison based on regions, the study also looks at differences of opinions based on property size.

Web-Based Training in the U.S Lodging Industry

Menu analysis is the gathering and processing of key pieces of information to make it more manageable and understand- able. Ultimately, menu analysis allows managers to make more informed decisions about prices, costs, and items to be included on a menu. The author discusses If labor as well as food casts need to be included in menu analysis and if managers need to categorize menu items differently when doing menu analysis based on customer eating patterns.

HDL-C and HDL-C/ApoA-I predict long-term progression of glycemia in established type 2 diabetes

OBJECTIVE: Low HDL cholesterol (HDL-C) and small HDL particle size may directly promote hyperglycemia. We evaluated associations of HDL-C, apolipoprotein A-I (apoA-I), and HDL-C/apoA-I with insulin secretion, insulin resistance, HbA1c, and long-term glycemic deterioration, reflected by initiation of pharmacologic glucose control.

RESEARCH DESIGN AND METHODS: The 5-year Fenofibrate Intervention and Event Lowering in Diabetes (FIELD) study followed 9,795 type 2 diabetic subjects. We calculated baseline associations of fasting HDL-C, apoA-I, and HDL-C/apoA-I with HbA1c and, in those not taking exogenous insulin (n = 8,271), with estimated β-cell function (homeostasis model assessment of β-cell function [HOMA-B]) and insulin resistance (HOMA-IR). Among the 2,608 subjects prescribed lifestyle only, Cox proportional hazards analysis evaluated associations of HDL-C, apoA-I, and HDL-C/apoA-I with subsequent initiation of oral hypoglycemic agents (OHAs) or insulin.

RESULTS: Adjusted for age and sex, baseline HDL-C, apoA-I, and HDL-C/apoA-I were inversely associated with HOMA-IR (r = -0.233, -0.134, and -0.230; all P < 0.001; n = 8,271) but not related to HbA1c (all P > 0.05; n = 9,795). ApoA-I was also inversely associated with HOMA-B (r = -0.063; P = 0.002; n = 8,271) adjusted for age, sex, and HOMA-IR. Prospectively, lower baseline HDL-C and HDL-C/apoA-I levels predicted greater uptake (per 1-SD lower: hazard ratio [HR] 1.13 [CI 1.07-1.19], P < 0.001; and HR 1.16 [CI 1.10-1.23], P < 0.001, respectively) and earlier uptake (median 12.9 and 24.0 months, respectively, for quartile 1 vs. quartile 4; both P < 0.01) of OHAs and insulin, with no difference in HbA1c thresholds for initiation (P = 0.87 and P = 0.81). Controlling for HOMA-IR and triglycerides lessened both associations, but HDL-C/apoA-I remained significant.

CONCLUSIONS: HDL-C, apoA-I, and HDL-C/apoA-I were associated with concurrent insulin resistance but not HbA1c. However, lower HDL-C and HDL-C/apoA-I predicted greater and earlier need for pharmacologic glucose control.

Faktorer som påverkar omvårdnaden av patienter från olika kulturer inom palliativ vård – en litteraturöversikt

När befolkningen i Sverige blir allt äldre kommer det bland annat leda till ett ökat sjukvårdsbehov. I genomsnitt har människor födda i länder långt från Norden sämre hälsa än personer födda inom Norden, de kan ha en annan kultur och tolka kommunikationen inom vården annorlunda. Syfte: Att beskriva faktorer som påverkar omvårdnaden av patienter och anhöriga från andra kulturer inom palliativ vård. Metod: En litteraturöversikt. Resultat: Är baserat på nio vetenskapliga artiklar. Det fanns kulturella skillnader i synen på allvarlig sjukdom och död. Inom flera kulturer är det tabubelagt att tala om döden och detta var sjuksköterskorna inom västerländsk kultur oerfarna vid, de var mer vana vid en rak och ärlig kommunikation med patienterna. Sjuksköterskorna hade kommunikationssvårigheter med patienter och närstående då de inte hade samma språk. Det var även svårt och tidskrävande att finna lämpliga tolkar som kunde hantera känsliga ämnen relaterat till palliativ vård. Konklusion: Sjuksköterskor upplevde att möten med patienter och anhöriga från andra kulturer ofta var svåra om de inte pratade samma språk. Trots detta upplevde sjuksköterskorna arbetet som tillfredsställande. Det framkom att för lite tid och stress var ett av problemen samt att behovet av tolkar var stort.

The Comfort of life depends upon Conversation – Puheen ja keskustelun ihanteet 1700-luvun englantilaisessa miehille ja naisille suunnatussa käytöskirjallisuudessa

Tutkielmassani tarkastellaan 1700-luvun englantilaisten käytösoppaiden antamia ohejeita ihanteellisesta kohteliaasta puheesta ja keskustelusta. Kohtelias käytöskulttuuri oli kehittynyt 1700-luvun kuluessa kaikenkattavaksi kulttuuriseksi ideologiaksi, jonka tärkein ilmaisumuoto oli keskustelu. Tutkielmassani pyrin selvittämään valitsemani neljän 1700-luvun englantilaisten käytösoppaan antamia ohjeita ihanteellisesta keskustelusta ja puheesta. Kaksi opasta on suunnattu miehille ja kaksi vastaavasti naisille. Tarkastelen naisille ja miehille suunnattujen 1700-luvun englantilaisten käytösoppaiden antamia ohjeita ihanteellisesta keskustelusta ja puheesta. Tärkeäksi näkökulmani kannalta muodostuu ajan kohtelias käytöskoodisto ja sen suhde puheeseen. Pohdin työssäni kohteliaisuuden suhdetta puheeseen ajan käytöskirjallisuuden kautta. Millaista kohtelias puhe niiden mukaan oli ja millaisia ideologisia sisältöjä sekä tavoitteita se sisälsi. Tuon esiin myös kohteliaisuuden suhdetta 1700-luvun englantilaisen yläluokan elämään ja sen vaikutukseen yhteiskunnassa. Pohdin erityisesti sitä millaiseksi käytösoppaat kuvasivat miesten ja naisten ihanteellista puhetta sekä keskustelua. Millaisina puhujina oppaat näkivät miehen ja naisen? Erosiko miesten ja naisten puhe jotenkin toisistaan? Käytösoppaiden tapaan nähdä sukupuolet vaikuttaa vahvasti 1700-luvun Englannissa vallinneet käsitykset sukupuolten rooleista ja sukupuolten eroista. Toisaalta kohtelias käytöskulttuuri antaa ihanteelliselle keskustelulle ja puheelle oman kehyksensä. 1700-luvun englantilaiset käytösoppaat korostavat kohteliaan puheen roolia ihmiselämän ja sosiaalisten suhteiden kannalta. Se oli tärkein keino, jolla eliitti pystyi omaa asemaansa perustelevat ja jonka avulla yhteisöön kuuluminen arvioidaan. Kohtelias puhe vaati harjoittajiltaan paljon ja se vaatikin harjaantumista. Käytösoppaat näkevät miehen ja naisen hyvin erilaisina keskustelijoina. Molemmat pyrkivät puheellaan miellyttämään muita ja olemaan osa kohteliasta yhteisöä, mutta miesten ja naisten saamat puhujanroolit ovat hyvin erilaisia. Ihanteellinen maskuliininen puhe henkilöityy kohteliaaseen herrasmieheen, joka kohtelee puheellaan kanssaihmisiä hyväntahtoisesti ja yhdenvertaisesti. Käytösoppaat rajaavat kohteliaan herrasmiehen ideaalin ulkopuolelle naismaisia ja moraalittomia käytösmalleja maskuliinisuuden sekä kohteliaisuuden suojelemiseksi. Naisille puolestaan käytösoppaat antavat rajoitetun roolin kohteliaassa yhteisössä, jonka toiminnot ohjautuvat tämän oman maineen, mutta myös miehisen kunnian ja tarpeiden kautta. Toisaalta naisten puhe on kohteliaan keskustelun kannalta myös tärkeää ja se saa erityisen roolin avioliitossa.

Maailma joka muuttuu elokuvallisessa prosessissa – Ranskalaisen päiväkirjan (1961) dokumentaarisuudesta

Tutkimuksen aiheena on Jean Rouchin ja Edgar Morinin ohjaama elokuva Ranskalainen päiväkirja vuodelta 1961 ja sen dokumentaarisuus. Tutkielma osoittaa, miten cinéma vérité -tyylisuunta on tapa, jolla dokumentaarisuus Ranskalaisessa päiväkirjassa toteutuu. Tutkin, minkälaisia uusia näkökulmia teos pystyy dokumentaarisuudellaan aikakauteensa avaamaan. Tyylisuunta-ajattelun lisäksi tarkastelen dokumentaarisuutta myös metodina ja elokuvallisena keinona. Ranskalaisen päiväkirjan ohella alkuperäislähteenä on dokumenttielokuva Un été + 50 (2011), jossa elokuvan tekijät ja henkilöt kertovat tutkittavan elokuvan taustoista 50 vuotta elokuvan ilmestymisen jälkeen. Ranskalainen päiväkirja sijoittuu 1950- ja 1960-lukujen vaihteeseen, jolloin Ranska oli muuttumassa. Suurvalta-asema oli menetetty, ja uutta hyvinvointivaltiota rakennettiin epäyhtenäisen ja vieraantuneen kansakunnan varaan. Kolonialismin jälkimainingeissa ranskalaisten oli määriteltävä suhteensa muun muassa käynnissä olleeseen Algerian sotaan. Autosta oli tullut kulutuskulttuurin symboli, mutta kehittyvä teknologia alkoi tarjota mahdollisuuksia myös elokuvalle ja sen ilmaisukeinojen laajentamiselle. Tutkielma selvittää, miten uusi elokuvateknologia, elokuvanteon representaatiot ja elokuvan välittämä sisältö kytkeytyvät yhteen ja muodostavat perustan Ranskalaisen päiväkirjan dokumentaarisuudelle. Elokuva antaa mahdollisuuden pohtia, miten teknologia ja eri aikakausien ajatuskehikot ovat toimineet vuorovaikutuksessa keskenään. Tyylisuunta-ajattelu kytkee elokuvan myös aikakauden filosofisiin ja elokuvateoreettisiin virtauksiin. Muutoksen mahdollisuudet, todellisuuden luonne ja paikka askarruttivat niin ikään filosofeja, sosiologeja ja muiden alojen taiteilijoita.