CTLs directed against HER2 specifically cross-react with HER3 and HER4

The human epidermal growth factor receptor 2 (HER2) has been targeted as a breast cancer-associated Ag by T cell-based immunotherapeutical strategies such as cancer vaccines and adoptive T cell transfer. The prerequisite for a successful T cell-based therapy is the induction of T cells capable of recognizing the HER2-expressing tumor cells. In this study, we generated human cytotoxic T cell clones directed against the HER2(369-377) epitope known to be naturally presented with HLA-A*0201. Those HER2-reactive CTLs, which were also tumor lytic, exhibited a similar lysis pattern dividing the targets in lysable and nonlysable tumor cells. Several HER2-expressing tumor cells became susceptible to CTL-mediated lysis after IFN-gamma treatment and, in parallel, up-regulated molecules of the Ag-presenting machinery, indicating that the tumor itself also contributes to the success of CTL-mediated killing. Some of the HER2(369-377)-reactive T cells specifically cross-reacted with the corresponding peptides derived from the family members HER3 and/or HER4 due to a high sequence homology. The epitopes HER3(356-364) and HER4(361-369) were endogenously processed and contributed to the susceptibility of cell lysis by HER cross-reacting CTLs. The principle of "double" or "triple targeting" the HER Ags by cross-reacting T cells will impact the further development of T cell-based therapies.     

Body Mass Index,Smoking and Hypertensive Disorders during Pregnancy: A Population Based Case-Control Study

While obesity is an indicated risk factor for hypertensive disorders of pregnancy, smoking during pregnancy has been shown to be inversely associated with the development of preeclampsia and gestational hypertension. The purpose of this study was to investigate the combined effects of high body mass index and smoking on hypertensive disorders during pregnancy. This was a case-control study based on national registers, nested within all pregnancies in Iceland 1989–2004, resulting in birth at the Landspitali University Hospital. Cases (n = 500) were matched 1:2 with women without a hypertensive diagnosis who gave birth in the same year. Body mass index (kg/m²) was based on height and weight at 10–15 weeks of pregnancy. We used logistic regression models to calculate odds ratios and corresponding 95% confidence intervals as measures of association, adjusting for potential confounders and tested for additive and multiplicative interactions of body mass index and smoking. Women’s body mass index during early pregnancy was positively associated with each hypertensive outcome. Compared with normal weight women, the multivariable adjusted odds ratio for any hypertensive disorder was 1.8 (95% confidence interval, 1.3–2.3) for overweight women and 3.1 (95% confidence interval, 2.2–4.3) for obese women. The odds ratio for any hypertensive disorder with obesity was 3.9 (95% confidence interval 1.8–8.6) among smokers and 3.0 (95% confidence interval 2.1–4.3) among non-smokers. The effect estimates for hypertensive disorders with high body mass index appeared more pronounced among smokers than non-smokers, although the observed difference was not statistically significant. Our findings may help elucidate the complicated interplay of these lifestyle-related factors with the hypertensive disorders during pregnancy.     

Picoplankton culture assessment using single strand conformation polymorphism and partial 18S sequencing

Environmental water samples were taken throughout 2001 fromfractioned water samples at the Helgoland time series site.The less than 3-µm fraction was inoculated into variousmedia. Serial dilutions from these inoculations produced a largenumber of rough cultures from which several hundred well-growingpicoplankton cultures were started. We established a combinationof crude DNA extraction, single strand conformation polymorphism(SSCP) fingerprinting and subsequent sequence analysis to screenthese large numbers of cultures, assess their purity/clonalityand determine their identity. Picoplankton species (i.e., cellssmaller than 3 µm) are difficult to distinguish becauseof their small size and the lack of morphological characters.Therefore, molecular techniques provide the most reliable methodto achieve their identification. Cultures were enriched forphotoautotroph cells, i.e., no carbon source was added, andcultures were grown in the light. From these cultures, crudeDNA was extracted, which was used for partial 18S polymerasechain reaction (PCR). On average, 50% of the cultures produceda PCR product. SSCP analysis of PCR products revealed the clonalityof a given culture. If clonal, then there was only a singleSSCP band; if not clonal, then there were multiple SSCP bands.Single SSCP bands were subsequently sequenced, and sequenceswere used to identify the culture. For this study, 300 cultureswere screened resulting in the identification of 63 potentiallyclonal cultures. These methods proved to be relatively easyto apply to assess the clonality and purity of the cultures.     

Polycystic nephropathy in slender lorises (Loris lydekkerianus)

In a colony of slender lorises, 20 deaths that occurred over a period of 11 years were investigated postmortem. Juvenile/adult polycystic nephropathy was observed in one newborn and 13 adult slender lorises. Although polycystic kidney disease (PKD) in humans and other animals is known to be inherited, it is not clear whether kidney alterations in slender lorises are genetically transmitted, stress related, or induced by microbiological influences.     

Effects of elevated parameters of subclinical ketosis on the immune system of dairy cows: in vivo and in vitro results

Using an established model in which subclinical ketosis is induced, the response of differential blood counts and levels of various haematological variables, including the inflammatory marker haptoglobin (Hp), were tested over the last six weeks of parturition until the 56th day post-partum in cows with lower or higher body condition scores (LBC and HBC, respectively; n = 9/group). Animals in the HBC group evidenced subclinical ketosis whereas LBC animals were metabolically healthy. For in vitro examination with ß-hydroxybutyrate (BHB) as a further stimulus, peripheral blood mononuclear cell (PBMC) counts of cows with and without subclinical ketosis (n = 5/group) were observed. Counts of leucocytes, granulocytes and lymphocytes (LY) peaked at day 1 post-partum in HBC cows, with a more marked increase in heifers. In subclinical ketosis LY count increased again, with significantly higher values in the HBC group. The red blood cell (RBC) profile was affected by parity (counts were higher in heifers). Hp showed a positive linear correlation with BHB and non-esterified fatty acids (NEFA; R² = 0.41). PBMC from cows that were not pre-stressed with subclinical ketosis were more sensitive to increasing levels of BHB in vitro, as evidenced by both their higher proliferative capability and increased release of nitric oxide (NO). In summary, cows with subclinical ketosis showed a heightened immune response compared with metabolically healthy individuals, based on increased LY counts, increasing stimulative properties of PBMC and a relationship between Hp and typically increased values of BHB and NEFA. Concentrations of BHB in vivo during subclinical ketosis did not alter the proliferative capability of bovine PBMC in vitro, which was first significantly decreased at a dosage of 5 mM BHB.     

Thermal acclimation,mitochondrial capacities and organ metabolic profiles in a reptile (<Emphasis Type=Italic>Alligator mississippiensis</Emphasis>)

Reptiles thermoregulate behaviourally, but change their preferred temperature and the optimal temperature for performance seasonally. We evaluated whether the digestive and locomotor systems of the alligator show parallel metabolic adjustments during thermal acclimation. To this end, we allowed juvenile alligators to grow under thermal conditions typical of winter and summer, providing them with seasonally appropriate basking opportunities. Although mean body temperatures of alligators in these groups differed by approximately 10°C, their growth and final anatomic status was equivalent. While hepatic mitochondria isolated from cold-acclimated alligators had higher oxidative capacities at 30°C than those from warm-acclimated alligators, the capacities did not differ at 20°C. Cold acclimation decreased maximal oxidative capacities of muscle mitochondria. For mitochondria from both organs and acclimation groups, palmitate increased oligomycin-inhibited respiration. GDP addition reduced palmitate-uncoupled rates more in liver mitochondria from warm- than cold-acclimated alligators. In muscle mitochondria, carboxyatractyloside significantly reduced palmitate-uncoupled rates. This effect was not changed by thermal acclimation. The aerobic capacity of liver, skeletal muscle and duodenum, as estimated by activities of cytochrome c oxidase (COX), increased with cold acclimation. At acclimation temperatures, the activities of COX and citrate synthase (CS) in these organs were equivalent. By measuring COX and CS in isolated mitochondria and tissue extracts, we estimated that cold acclimation did not change the mitochondrial content in liver, but increased that of muscle. The thermal compensation of growth rates and of the aerobic capacity of the locomotor and digestive systems suggests that alligators optimised metabolic processes for the seasonally altered, preferred body temperature. The precision of this compensatory response exceeds that typically shown by aquatic ectotherms whose body temperatures are at the mercy of their habitat.     

Terahertz Electromagnetic Fields (0.106 THz) Do Not Induce Manifest Genomic Damage In Vitro

Terahertz electromagnetic fields are non-ionizing electromagnetic fields in the frequency range from 0.1 to 10 THz. Potential applications of these electromagnetic fields include the whole body scanners, which currently apply millimeter waves just below the terahertz range, but future scanners will use higher frequencies in the terahertz range. These and other applications will bring along human exposure to these fields. Up to now, only a limited number of investigations on biological effects of terahertz electromagnetic fields have been performed. Therefore, research is strongly needed to enable reliable risk assessment.Cells were exposed for 2 h, 8 h, and 24 h with different power intensities ranging from 0.04 mW/cm² to 2 mW/cm², representing levels below, at, and above current safety limits. Genomic damage on the chromosomal level was measured as micronucleus formation. DNA strand breaks and alkali-labile sites were quantified with the comet assay. No DNA strand breaks or alkali-labile sites were observed as a consequence of exposure to terahertz electromagnetic fields in the comet assay. The fields did not cause chromosomal damage in the form of micronucleus induction.     

Endocytosis and intracellular processing of BODIPY-sphingomyelin by murine CATH.a neurons

Neuronal sphingolipids (SL) play important roles during axonal extension, neurotrophic receptor signaling and neurotransmitter release. Many of these signaling pathways depend on the presence of specialized membrane microdomains termed lipid rafts. Sphingomyelin (SM), one of the main raft constituents, can be formed de novo or supplied from exogenous sources. The present study aimed to characterize fluorescently-labeled SL turnover in a murine neuronal cell line (CATH.a). Our results demonstrate that at 4 °C exogenously added BODIPY-SM accumulates exclusively at the plasma membrane. Treatment of cells with bacterial sphingomyelinase (SMase) and back-exchange experiments revealed that 55–67% of BODIPY-SM resides in the outer leaflet of the plasma membrane. Endocytosis of BODIPY-SM occurs via caveolae with part of internalized BODIPY-fluorescence ending up in the Golgi and the ER. Following endocytosis BODIPY-SM undergoes hydrolysis, a reaction substantially faster than BODIPY-SM synthesis from BODIPY-ceramide. RNAi demonstrated that both, acid (a)SMase and neutral (n)SMases contribute to BODIPY-SM hydrolysis. Finally, high-density lipoprotein (HDL)-associated BODIPY-SM was efficiently taken up by CATH.a cells. Our findings indicate that endocytosis of exogenous SM occurs almost exclusively via caveolin-dependent pathways, that both, a- and nSMases equally contribute to neuronal SM turnover and that HDL-like particles might represent physiological SM carriers/donors in the brain.