Impact of HLA-B alleles, epitope binding affinity, functional avidity, and viral coinfection on the immunodominance of virus-specific CTL responses

Immunodominance is variably used to describe either the most frequently detectable response among tested individuals or the strongest response within a single individual, yet factors determining either inter- or intraindividual immunodominance are still poorly understood. More than 90 individuals were tested against 184 HIV- and 92 EBV-derived, previously defined CTL epitopes. The data show that HLA-B-restricted epitopes were significantly more frequently recognized than HLA-A- or HLA-C-restricted epitopes. HLA-B-restricted epitopes also induced responses of higher magnitude than did either HLA-A- or HLA-C-restricted epitopes, although this comparison only reached statistical significance for EBV epitopes. For both viruses, the magnitude and frequency of recognition were correlated with each other, but not with the epitope binding affinity to the restricting HLA allele. The presence or absence of HIV coinfection did not impact EBV epitope immunodominance patterns significantly. Peptide titration studies showed that the magnitude of responses was associated with high functional avidity, requiring low concentration of cognate peptide to respond in in vitro assays. The data support the important role of HLA-B alleles in antiviral immunity and afford a better understanding of the factors contributing to inter- and intraindividual immunodominance.     

Determination of the period for establishment of a liver network echogenic pattern in Schistosoma japonicum infection

Schistosomiasis is caused by infection with Schistosoma haematobium, S. mansoni, S. japonicum, or S. mekongi. S. japonicum infection results in liver cirrhosis at the final stage. A "network" (NW) echogenic pattern on hepatic ultrasonography appears to be specific to S. japonicum infection. The principal aim of the present study was to determine the exact year(s) or even month(s) required for the establishment of the liver NW echogenic pattern from the initial infection in young patients with schistosomiasis japonica since there are few data on this important point. We conducted yearly ultrasonographic, serologic, coprologic, and physical examinations of schistosomiasis patients in the Philippines from 1996 up to the present. During that period, the total number of patients examined was approximately 2,000, among whom we selected 2 patients for determination of the duration required for NW establishment, when they were 10 years old. Although the exact time of initial exposure to schistosomes cannot be determined, the duration for the establishment of NW was definitively confirmed in patient no. 1 to be between 19-24 months based on the results of serologic and coprologic examinations. For patient no. 2, the circumstantial evidence suggested that the establishment of a NW might require 5 to 6 years at maximum. To the best of our knowledge, this is the first evidence-based report on the determination of the period required for the establishment of a liver NW echogenic pattern in S. japonicum infection in the Philippines.     

Abundance of Lutzomyia longipalpis (Diptera: Psychodidae: Phlebotominae) and urban transmission of visceral leishmaniasis in Campo Grande, state of Mato Grosso do Sul, Brazil

The outspread and urbanization of visceral leishmaniasis (VL) in Campo Grande, state of Mato Grosso do Sul, lead us to undertake the present study over diversity and abundance of sand flies in the urban area to compare with previous search carried out during 1999/2000, before the identification of the disease in the human population.The captures were carried out with automatic light traps, weekly, from February 2004 to February 2005 on three sites including a forested area (Zé Pereira), two peridomicilies (shelters of domestic animals and cultivation areas), and intradomicilie. In the present study 110 collections were obtained during 13 months for 1320 h of collections, resulting in 5004 specimens, 3649 males and 1355 females belonging to the 20 following species: Brumptomyia avellari, Brumptomyia sp., Bichromomyia flaviscutellata, Evandromyia lenti, E. termitophila, E. cortelezzii, E. borrouli, Lutzomyia sp., L. longipalpis, Micropygomyia quinquefer, N. antunesi, N. whitmani, Pintomyia christenseni, Pi. damascenoi, Psathyromyia aragaoi, Ps. campograndensis, Ps. hermanlenti, Ps. shannoni, Pychodopygus claustrei, and Sciopemyia sordellii. L. longipalpis was the most abundant species in the anthropic environment with 92.22% of the captures. This shows an increase of sixty times in the density of L. longipalpis compared to the last sand fly evaluation in 1999/2000. The high density of L. longipalpis in Campo Grande is the main factor of risk in transmission of the disease to human in the urban area. The capture of N. antunesi, typical specie from Amazonian region, in Mato Grosso do Sul is reported for the first time.     

Surface roughness of denture base and reline materials after disinfection by immersion in chlorhexidine or microwave irradiation

doi: 10.1111/j.1741‐2358.2011.00484.x
Surface roughness of denture base and reline materials after disinfection by immersion in chlorhexidine or microwave irradiation Background: This study evaluated the effect of disinfection by immersion and microwave irradiation on the roughness of one denture base resin (Lucitone‐L) and five relining materials, three hard (Tokuyama Rebase II‐TR, New Truliner‐NT, Ufigel Hard‐UH) and two resilient (Trusoft‐T, Sofreliner‐S). Methods: Fifty specimens were made and divided into groups: CL2 specimens were brushed with 4% chlorhexidine (1 min), immersed in the same solution (10 min) and immersed in water (3 min); MW2 specimens were immersed in water and microwave irradiated (650W; 6 min); CL2 and MW2 specimens were disinfected twice; CL7 and MW7 specimens were submitted to seven cycles using chlorhexidine or microwave irradiation, respectively; W specimens were not disinfected and remained in water (37°C; 7 days). Results: Results were statistically analysed (p = 0.05) and revealed that, at baseline, the highest mean value was observed for T (p < 0.001). Material NT showed increase in roughness after the first (p = 0.003), second (p = 0.001), seventh (p = 0.000) cycles of microwave disinfection and after 7 days of immersion in water (p = 0.033). Conclusions: Resilient liner S presented significant increase in roughness after the second cycle of disinfection with chlorhexidine (p = 0.003). Material T exhibited significantly decreased roughness in group W (p = 0.010), while microwaving produced severe alterations on its surface.     

Mutations in SRCAP, encoding SNF2-related CREBBP activator protein, cause Floating-Harbor syndrome

Floating-Harbor syndrome (FHS) is a rare condition characterized by short stature, delayed osseous maturation, expressive-language deficits, and a distinctive facial appearance. Occurrence is generally sporadic, although parent-to-child transmission has been reported on occasion. Employing whole-exome sequencing, we identified heterozygous truncating mutations in SRCAP in five unrelated individuals with sporadic FHS. Sanger sequencing identified mutations in SRCAP in eight more affected persons. Mutations were de novo in all six instances in which parental DNA was available. SRCAP is an SNF2-related chromatin-remodeling factor that serves as a coactivator for CREB-binding protein (CREBBP, better known as CBP, the major cause of Rubinstein-Taybi syndrome [RTS]). Five SRCAP mutations, two of which are recurrent, were identified; all are tightly clustered within a small (111 codon) region of the final exon. These mutations are predicted to abolish three C-terminal AT-hook DNA-binding motifs while leaving the CBP-binding and ATPase domains intact. Our findings show that SRCAP mutations are the major cause of FHS and offer an explanation for the clinical overlap between FHS and RTS.     

Haploinsufficiency of a spliceosomal GTPase encoded by EFTUD2 causes mandibulofacial dysostosis with microcephaly

Mandibulofacial dysostosis with microcephaly (MFDM) is a rare sporadic syndrome comprising craniofacial malformations, microcephaly, developmental delay, and a recognizable dysmorphic appearance. Major sequelae, including choanal atresia, sensorineural hearing loss, and cleft palate, each occur in a significant proportion of affected individuals. We present detailed clinical findings in 12 unrelated individuals with MFDM; these 12 individuals compose the largest reported cohort to date. To define the etiology of MFDM, we employed whole-exome sequencing of four unrelated affected individuals and identified heterozygous mutations or deletions of EFTUD2 in all four. Validation studies of eight additional individuals with MFDM demonstrated causative EFTUD2 mutations in all affected individuals tested. A range of EFTUD2-mutation types, including null alleles and frameshifts, is seen in MFDM, consistent with haploinsufficiency; segregation is de novo in all cases assessed to date. U5-116kD, the protein encoded by EFTUD2, is a highly conserved spliceosomal GTPase with a central regulatory role in catalytic splicing and post-splicing-complex disassembly. MFDM is the first multiple-malformation syndrome attributed to a defect of the major spliceosome. Our findings significantly extend the range of reported spliceosomal phenotypes in humans and pave the way for further investigation in related conditions such as Treacher Collins syndrome.     

Sex‐biased resource allocation in ovo in a sexually size‐dimorphic species

Sex‐biased resource allocation in eggs is increasingly recognized as one strategy oviparous mothers can employ to invest differentially in one sex, depending on nutritional requirements. Previous studies have used egg size as an index of nutrient allocation, but few have examined egg contents directly. We used molecular sexing of early‐stage ring‐billed gull Larus delawarensis embryos, a species with sexual size dimorphism, to test whether sex‐specific nutrient allocation occurs in ovo. Despite no sex difference in size, eggs with male embryos contained more albumen, while eggs with female embryos contained more yolk, lipid and non‐lipid (protein and carbohydrate). It is unclear why such sex‐biased resource allocation in ovo is utilized by ring‐billed gulls. However, our data indicate that a cursive examination of egg mass or size may not necessarily reflect nutrient allocation strategies mothers use in ovo, and that sex‐biased investment in ovo may be more widespread than currently appreciated.     

Satellite 2 demethylation induced by 5-azacytidine is associated with missegregation of chromosomes 1 and 16 in human somatic cells

Satellite sequences are an important part of the pericentromeric regions in mammalian genomes; they play a relevant role in chromosome stability and DNA hypomethylation of these sequences has been reported in ICF syndrome and in some cancers that are closely associated with chromosomal abnormalities. Epigenetic modifications of satellite sequences and their consequences have not been extensively studied in human cells. In the present work, we evaluated satellite 2 methylation patterns in human lymphocytes exposed to 5-azacytidine (5-azaC) and assessed the relationship between these patterns and chromosome missegregation. Human lymphocytes were exposed to 10μM 5-azaC for 24, 48, and 72h. Segregation errors were evaluated in binucleate cells using FISH against pericentromeric regions of chromosomes 1, 9, and 16. DNA methylation patterns were evaluated by immunodetection, and by bisulfite plus urea conversion and sequencing. We have identified that 5-azaC induced missegregation of chromosomes 1 and 16, which have highly methylated satellite 2, after 72h of exposure. Chromosome methylation patterns showed a notable decrease in pericentromeric methylation. Bisulfite conversion and sequencing analysis demonstrated demethylation of satellite 2 associated to 5-azaC exposure, principally after 72h of treatment. This change occurred in a non-specific pattern. Our study demonstrates an association between loss of satellite 2 DNA methylation and chromosome loss in human lymphocytes.     

An MRM-based workflow for quantifying cardiac mitochondrial protein phosphorylation in murine and human tissue

The regulation of mitochondrial function is essential for cardiomyocyte adaptation to cellular stress. While it has long been understood that phosphorylation regulates flux through metabolic pathways, novel phosphorylation sites are continually being discovered in all functionally distinct areas of the mitochondrial proteome. Extracting biologically meaningful information from these phosphorylation sites requires an adaptable, sensitive, specific and robust method for their quantification. Here we report a multiple reaction monitoring-based mass spectrometric workflow for quantifying site-specific phosphorylation of mitochondrial proteins. Specifically, chromatographic and mass spectrometric conditions for 68 transitions derived from 23 murine and human phosphopeptides, and their corresponding unmodified peptides, were optimized. These methods enabled the quantification of endogenous phosphopeptides from the outer mitochondrial membrane protein VDAC, and the inner membrane proteins ANT and ETC complexes I, III and V. The development of this quantitative workflow is a pivotal step for advancing our knowledge and understanding of the regulatory effects of mitochondrial protein phosphorylation in cardiac physiology and pathophysiology. This article is part of a Special Issue entitled: Translational Proteomics.