Multiple mechanisms compensate to enhance tumor-protective CD8(+) T cell response in the long-term despite poor CD8(+) T cell priming initially: comparison between an acute versus a chronic intracellular bacterium expressing a model antigen

We evaluated CD8(+) T cell responses against the dominant CTL epitope, OVA(257-264), expressed by an acute (Listeria monocytogenes (LM) OVA) vs a chronic pathogen (Mycobacterium bovis bacillus Calmette-Guérin (BCG) OVA) to reveal the influence on CD8(+) T cell memory and consequent protection against a challenge with OVA-expressing tumor cells. Infection with lower doses of both pathogens resulted in stronger bacterial growth but weaker T cell memory indicating that memory correlates with pathogen dose but not with bacterial expansion. The CD8(+) T cell response induced by LM-OVA was helper T cell-independent and was characterized by a rapid effector response followed by a rapid, but massive, attrition. In contrast, BCG-OVA induced a delayed and weak response that was compensated for by a longer effector phase and reduced attrition. This response was partly dependent on CD4(+) T cells. CD8(+) T cell response induced by BCG-OVA, but not LM-OVA, was highly dependent on pathogen persistence to compensate for the weak initial CD8(+) T cell priming. Despite a stronger initial T cell response with LM-OVA, BCG-OVA provided more effective tumor (B16OVA) control at both local and distal sites due to the induction of a persistently activated acquired, and a more potent innate, immunity.     

Quantitation of platelet adhesion to Ti and DLC-coated Ti in vitro using (125)I-labeled platelets

Measurement of platelet adhesion in vitro is a good indicator of its reactivity to implant devices in vivo. Platelets were labeled with I-125 without affecting its normal morphology and function and the labeled platelets were suspended in platelet poor plasma and exposed to Ti and diamond like carbon-coated (DLC) Ti discs, under dynamic conditions, using a parallel plate flow chamber. The test materials were washed, dried, exposed to a phosphor screen and scanned to get the images. The number of platelets that adhered to Ti was found to be higher than those that adhered to DLC coated Ti sample, irrespective of the shear stress which was varied between 2 and 16 dynes/cm(2).     

The Chediak-Higashi protein interacts with SNARE complex and signal transduction proteins

BACKGROUND:Chediak-Higashi syndrome (CHS) is an inherited immunodeficiency disease characterized by giant lysosomes and impaired leukocyte degranulation. CHS results from mutations in the lysosomal trafficking regulator (LYST) gene, which encodes a 425-kD cytoplasmic protein of unknown function. The goal of this study was to identify proteins that interact with LYST as a first step in understanding how LYST modulates lysosomal exocytosis. MATERIALS AND METHODS: Fourteen cDNA fragments, covering the entire coding domain of LYST, were used as baits to screen five human cDNA libraries by a yeast two-hybrid method, modified to allow screening in the activation and the binding domain, three selectable markers, and more stringent confirmation procedures. Five of the interactions were confirmed by an in vitro binding assay. RESULTS: Twenty-one proteins that interact with LYST were identified in yeast two-hybrid screens. Four interactions, confirmed directly, were with proteins important in vesicular transport and signal transduction (the SNARE-complex protein HRS, 14-3-3, and casein kinase II). CONCLUSIONS:On the basis of protein interactions, LYST appears to function as an adapter protein that may juxtapose proteins that mediate intracellular membrane fusion reactions. The pathologic manifestations observed in CHS patients and in mice with the homologous mutation beige suggest that understanding the role of LYST may be relevant to the treatment of not only CHS but also of diseases such as asthma, urticaria, and lupus, as well as to the molecular dissection of the CHS-associated cancer predisposition.     

Evaluation of genetic diversity among Pseudomonas citronellolis strains isolated from oily sludge-contaminated sites

The diversity among a set of bacterial strains that have the capacity to degrade total petroleum hydrocarbons (TPH) in soil contaminated with oily sludge (hazardous hydrocarbon waste from oil refineries) was determined. TPH is composed of alkane, aromatics, nitrogen-, sulfur-, and oxygen-containing compound, and asphaltene fractions of crude oil. The 150 bacterial isolates which could degrade TPH were isolated from soil samples obtained from diverse geoclimatic regions of India. All the isolates were biochemically characterized and identified with a Biolog microbial identification system and by 16S rDNA sequencing. Pseudomonas citronellolis predominated among the 150 isolates obtained from six different geographically diverse samplings. Of the isolates, 29 strains of P. citronellolis were selected for evaluating their genetic diversity. This was performed by molecular typing with repetitive sequence (Rep)-based PCR with primer sets ERIC (enterobacterial repetitive intergenic consensus), REP (repetitive extragenic palindromes), and BOXAIR and PCR-based ribotyping. Strain-specific and unique genotypic fingerprints were distinguished by these molecular typing strategies. The 29 strains of P. citronellolis were separated into 12 distinguishable genotypic groups by Rep-PCR and into seven genomic patterns by PCR-based ribotyping. The genetic diversity of the strains was related to the different geoclimatic isolation sites, type of oily sludge, and age of contamination of the sites. These results indicate that a combination of Rep-PCR fingerprinting and PCR-based ribotyping can be used as a high-resolution genomic fingerprinting method for elucidating intraspecies diversity among strains of P. citronellolis.     

Transforming growth factor-beta: a neuroprotective factor in cerebral ischemia

Transforming growth factor-β (TGF-β) has diverse and multiple roles throughout the body. This review focuses on the evidence supporting its functions in the central nervous system, with a particular emphasis on its purported role in cerebral ischemia. Numerous studies have documented that TGF-β1 levels are enhanced in the brain following cerebral ischemia. As evidence that such an upregulation is beneficial, agonist studies have demonstrated that TGF-β1 reduces neuronal cell death and infarct size following middle cerebral artery occlusion (MCAO), while conversely, antagonist studies have shown increased neuronal cell death and infarct size after MCAO. These studies suggest that TGF-β1 has a neuroprotective role in cerebral ischemia. Recent work with adenoviral-mediated overexpression of TGF-β1 in vivo in mice has further implicated a neuroprotective role for TGF-β1 in cerebral ischemia, as evidenced by a reduction in neuronal cell death, infarct size, and neurological outcome. Additionally, numerous in vitro studies have documented the neuroprotective ability of TGF-β1 in neurons from a variety of species, including rats, mice, chicks, and humans. Of significant interest, TGF-β1 was shown to be protective against a wide variety of death-inducing agents/insults, including hypoxia/ischemia, glutamate excitotoxicity, β-amyloid, oxidative damage, and human immunodeficiency virus. The mechanism of TGF-β1-mediated neuroprotection remains to be resolved, but early evidence suggests that TGF-β1 regulates the expression and ratio of apoptotic (Bad) and antiapoptotic proteins (Bcl-2, Bcl-x₁), creating an environment favorable for cell survival of death-inducing insults. Taken as a whole, these results suggest that TGF-β1 is an important neuroprotective factor that can reduce damage from a wide-array of death-inducing agents/insults in vitro, as well as exert protection of the brain during cerebral ischemia. The authors’ research is supported by research grants (HD-28964 and AG-17186 to DWB) from the National Institutes of Health, NICHD, and NIA.     

Smoking-gender interaction and risk for rheumatoid arthritis

The present case-control study was conducted to investigate the relationship between smoking and rheumatoid arthritis, and to investigate formally the interaction between sex, smoking, and risk for developing rheumatoid arthritis. The study was performed in the Central District of Finland. Cases were patients with rheumatoid arthritis and the control group was a random sample of the general population. Logistic regression models were used to evaluate the effect of smoking on risk for rheumatoid arthritis, after adjusting for the effects of age, education, body mass index, and indices of general health and pain. Overall, 1095 patients with rheumatoid arthritis and 1530 control individuals were included. Patients were older, less well educated, more disabled, and had poorer levels of general health as compared with control individuals (all P < 0.01). Preliminary analyses revealed the presence of substantial statistical interaction between smoking and sex (P < 0.001). In separate multivariable analyses, past history of smoking was associated with increased risk for rheumatoid arthritis overall in men (odds ratio 2.0, 95% confidence interval 1.2-3.2) but not in women. Among men, this effect was seen only for rheumatoid factor-positive rheumatoid arthritis. There were significant interactions between smoking and age among women but not among men. We conclude that sex is a biologic effect modifier in the association between smoking and rheumatoid arthritis. The role of menopause in the etiology of rheumatoid arthritis merits further research.     

Changes in NMR relaxation times in soybean and wheat seeds equilibrated at different temperatures and relative humidity

The relationship between equilibration injury and equilibration dependence of the transverse relaxation time (T2) measurements was examined using NMR in two different seed species (sensitive-soybean and tolerant-wheat) differing in their sensitivity to seed equilibration conditions. The T2 values of both seed species declined with high temperature (45 degrees C) and low RH (5.5-1%) and, also with high temperature (45 degrees C) and high RH (74.5-100%) conditions. A comparison of injury based on electrolyte leakage, seed germination percentage and T2 indicated that membrane permeability increased both at high temperature (45 degrees C) and low RH (5.5-1%) and high temperature (45 degrees C) and high RH (74.5-100%) seed equilibration conditions. There was an increase in T2 until 11.5% and 5.5% RH in soybean and wheat species respectively, followed by a decline. Loss of seed viability during equilibration at very low RH (5.5-1%) at 45 degrees C, and similarly at high RH (74.5-100%) at 45 degrees C indicates that the changes in T2 are probably due to the loss of membrane injury.     

An empirical relationship between rotational correlation time and solvent accessible surface area

Structure–dynamics interrelationships are important in understanding protein function. We have explored the empirical relationship between rotational correlation times (_c and the solvent accessible surface areas (SASA) of 75 proteins with known structures. The theoretical correlation between SASA and _c through the equation SASA = K_rc ^(2/3) is also considered. SASA was determined from the structure, _c ^calc was determined from diffusion tensor calculations, and _c ^expt was determined from NMR backbone¹³ C or ¹⁵N relaxation rate measurements. The theoretical and experimental values of _c correlate with SASA with regression analyses values of K_r as 1696 and 1896 m²s^-(2/3), respectively, and with corresponding correlation coefficients of 0.92 and 0.70.     

Screening and selection of media components for lactic acid production using Plackett–Burman design

Plackett–Burman design was used to efficiently select important media components influencing lactic acid production in a two step screening procedure. A total of 36 screening experiments were conducted for studying the effect of various media components such as carbon and nitrogen (simple and complex) sources, minerals/buffering agents and a specific inducer for the production of lactic acid by Lactobacillus plantarum NCIM 2084. The eleven ingredients chosen after the first screening experiments were further screened by a Plackett-Burman design consisting of 12 experiments. Liquefied starch, wheat bran extract, ammonium nitrate, manganese sulphate and sodium acetate were chosen as promising ingredients for further optimisation studies. The highest yield of 41.9?g/l of lactic acid was obtained at the end of 24 hours of fermentation which corresponded to 90% conversion, on the basis of sugar supplied.