Local delivery of AAV2-CTLA4IgG decreases sialadenitis and improves gland function in the C57BL/6.NOD-Aec1Aec2 mouse model of Sjögren's syndrome

Introduction

Cytotoxic T-lymphocyte antigen 4 (CTLA-4) is a key negative costimulatory molecule that displays a wide range of anti-inflammatory properties and is currently approved to treat rheumatoid arthritis as a recombinant fusion protein (CTLA4IgG). To better understand the role of CTLA4IgG in primary Sjögren's syndrome (pSS), we generated a recombinant adeno-associated virus vector serotype 2 (AAV2) expressing a chimera of mouse CTLA-4 fused with a human immunoglobulin (AAV2-CTLA4IgG) and observed the effect of this molecule in C57BL/6.NOD-Aec1Aec2 mice, an animal model of pSS.

Methods

A recombinant adeno-associated virus-2 (AAV-2) vector was constructed encoding a CTLA4IgG fusion protein. The AAV2-CTLA4IgG vector and an AAV2 control vector encoding beta galactosidase (LacZ) were administered by retrograde cannulation of the submandibular glands of C57BL/6.NOD-Aec1Aec2 mice. Protein expression was measured by ELISA and salivary glands were assessed for inflammation and activity.

Results

Recombinant CTLA4IgG blocked B7 expression on macrophages in vitro. In vivo, localized expression of CTLA4IgG in the salivary glands of C57BL/6.NOD-Aec1Aec2 mice inhibited the loss of salivary gland activity and decreased T and B cell infiltration as well as dendritic cells and macrophages in the glands compared with control mice. In addition a decrease in several proinflammatory cytokines and an increase in transforming growth factor beta-1 (TGF-β1) expression were also observed.

Conclusions

These data suggest expression of CTLA4IgG in the salivary gland can decrease the inflammation and improve the xerostomia reported in these mice.     

Thermal Reaction Norms and the Scale of Temperature Variation: Latitudinal Vulnerability of Intertidal Nacellid Limpets to Climate Change

The thermal reaction norms of 4 closely related intertidal Nacellid limpets, Antarctic (Nacella concinna), New Zealand (Cellana ornata), Australia (C. tramoserica) and Singapore (C. radiata), were compared across environments with different temperature magnitude, variability and predictability, to test their relative vulnerability to different scales of climate warming. Lethal limits were measured alongside a newly developed metric of “duration tenacity”, which was tested at different temperatures to calculate the thermal reaction norm of limpet adductor muscle fatigue. Except in C. tramoserica which had a wide optimum range with two break points, duration tenacity did not follow a typical aerobic capacity curve but was best described by a single break point at an optimum temperature. Thermal reaction norms were shifted to warmer temperatures in warmer environments; the optimum temperature for tenacity (T_opt) increased from 1.0°C (N. concinna) to 14.3°C (C. ornata) to 18.0°C (an average for the optimum range of C. tramoserica) to 27.6°C (C. radiata). The temperature limits for duration tenacity of the 4 species were most consistently correlated with both maximum sea surface temperature and summer maximum in situ habitat logger temperature. Tropical C. radiata, which lives in the least variable and most predictable environment, generally had the lowest warming tolerance and thermal safety margin (WT and TSM; respectively the thermal buffer of CT_max and T_opt over habitat temperature). However, the two temperate species, C. ornata and C. tramoserica, which live in a variable and seasonally unpredictable microhabitat, had the lowest TSM relative to in situ logger temperature. N. concinna which lives in the most variable, but seasonally predictable microhabitat, generally had the highest TSMs. Intertidal animals live at the highly variable interface between terrestrial and marine biomes and even small changes in the magnitude and predictability of their environment could markedly influence their future distributions.     

Iceberg Scour and Shell Damage in the Antarctic Bivalve Laternula elliptica

We document differences in shell damage and shell thickness in a bivalve mollusc (Laternula elliptica) from seven sites around Antarctica with differing exposures to ice movement. These range from 60% of the sea bed impacted by ice per year (Hangar Cove, Antarctic Peninsula) to those protected by virtually permanent sea ice cover (McMurdo Sound). Patterns of shell damage consistent with blunt force trauma were observed in populations where ice scour frequently occurs; damage repair frequencies and the thickness of shells correlated positively with the frequency of iceberg scour at the different sites with the highest repair rates and thicker shells at Hangar Cove (74.2% of animals damaged) compared to the other less impacted sites (less than 10% at McMurdo Sound). Genetic analysis of population structure using Amplified Fragment Length Polymorphisms (AFLPs) revealed no genetic differences between the two sites showing the greatest difference in shell morphology and repair rates. Taken together, our results suggest that L. elliptica exhibits considerable phenotypic plasticity in response to geographic variation in physical disturbance.     

Obstructed labor and caesarean delivery: the cost and benefit of surgical intervention

Background

Although advances in the reduction of maternal mortality have been made, up to 273,000 women will die this year from obstetric etiologies. Obstructed labor (OL), most commonly treated with Caesarean delivery, has been identified as a major contributor to global maternal morbidity and mortality. We used economic and epidemiological modeling to estimate the cost per disability-adjusted life-year (DALY) averted and benefit-cost ratio of treating OL with Caesarean delivery for 49 countries identified as providing an insufficient number of Caesarean deliveries to meet demand.

Methods and Findings

Using publicly available data and explicit economic assumptions, we estimated that the cost per DALY (3,0,0) averted for providing Caesarean delivery for OL ranged widely, from $251 per DALY averted in Madagascar to $3,462 in Oman. The median cost per DALY averted was $304. Benefit-cost ratios also varied, from 0.6 in Zimbabwe to 69.9 in Gabon. The median benefit-cost ratio calculated was 6.0. The main limitation of this study is an assumption that lack of surgical capacity is the main factor responsible for DALYs from OL.

Conclusions

Using the World Health Organization''s cost-effectiveness standards, investing in Caesarean delivery can be considered “highly cost-effective” for 48 of the 49 countries included in this study. Furthermore, in 46 of the 49 included countries, the benefit-cost ratio was greater than 1.0, implying that investment in Caesarean delivery is a viable economic proposition. While Caesarean delivery alone is not sufficient for combating OL, it is necessary, cost-effective by WHO standards, and ultimately economically favorable in the vast majority of countries included in this study.     

Selection, phenotyping and identification of Acid and hydrogen peroxide producing bacteria from vaginal samples of canadian and East african women

The common but poorly understood condition known as bacterial vaginosis (BV) increases vulnerability to HIV infection and is associated with the absence of H(2)O(2)-producing Lactobacillus. Vaginal lactic acid bacteria (LAB) produce anti-HIV factors such as organic acids and hydrogen peroxide (H(2)O(2)), and may bind and inactivate HIV particles during scavenging of mannose. These factors define potential criteria for initial selection of candidate probiotics to block heterosexual transmission of HIV. Therefore, the primary goal of this study was to characterize acid production on mannose and H(2)O(2) production in vaginal isolates from Canadian adolescents (192 isolates, 16 individuals) and commercial sex workers in Nairobi, Kenya (576 isolates, 96 individuals). Selection of isolates from H(2)O(2)-detecting media suggested an idiosyncratic individual-level profile and extensive phenotypic diversity, including the identification of a subset of "double-strong" acid- and H(2)O(2)-producers with phenotypes similar to well-characterized probiotic strains. Molecular fingerprinting of all isolates by capillary electrophoresis of 16S-23S rRNA interspacer amplicons was coupled with chaperonin-60 universal target (cpn60 UT) sequencing in a subset, tentatively identifying 96% of isolates although only 19% were sequenced. Most isolates belonged to Lactobacillus, Streptococcus, Bifidobacterium or Gardnerella, with a total of 37 species in 15 genera, as well as 5 potentially novel organisms, identified in this study. This sensitivity was likely enhanced by phenotype-based selection on two chromogenic media formulations. Identification of double-strong isolates may provide a rational basis for selection and further characterization of vaginal probiotics, with potential application as part of HIV prevention initiatives in western Canada and East Africa.     

Mechanism of foreign DNA selection in a bacterial adaptive immune system

In bacterial and archaeal CRISPR immune pathways, DNA sequences from invading bacteriophage or plasmids are integrated into CRISPR loci within the host genome, conferring immunity against subsequent infections. The ribonucleoprotein complex Cascade utilizes RNAs generated from these loci to target complementary "nonself" DNA sequences for destruction, while avoiding binding to "self" sequences within the CRISPR locus. Here we show that CasA, the largest protein subunit of Cascade, is required for nonself target recognition and binding. Combining a 2.3 ? crystal structure of CasA with cryo-EM structures of Cascade, we have identified a loop that is required for viral defense. This loop contacts a conserved three base pair motif that is required for nonself target selection. Our data suggest a model in which the CasA loop scans DNA for this short motif prior to target destabilization and binding, maximizing the efficiency of DNA surveillance by Cascade.