The same ELA class II risk factors confer equine insect bite hypersensitivity in two distinct populations

Insect bite hypersensitivity (IBH) is a chronic allergic dermatitis common in horses. Affected horses mainly react against antigens present in the saliva from the biting midges, Culicoides ssp, and occasionally black flies, Simulium ssp. Because of this insect dependency, the disease is clearly seasonal and prevalence varies between geographical locations. For two distinct horse breeds, we genotyped four microsatellite markers positioned within the MHC class II region and sequenced the highly polymorphic exons two from DRA and DRB3, respectively. Initially, 94 IBH-affected and 93 unaffected Swedish born Icelandic horses were tested for genetic association. These horses had previously been genotyped on the Illumina Equine SNP50 BeadChip, which made it possible to ensure that our study did not suffer from the effects of stratification. The second population consisted of 106 unaffected and 80 IBH-affected Exmoor ponies. We show that variants in the MHC class II region are associated with disease susceptibility (p (raw)?=?2.34?×?10(-5)), with the same allele (COR112:274) associated in two separate populations. In addition, we combined microsatellite and sequencing data in order to investigate the pattern of homozygosity and show that homozygosity across the entire MHC class II region is associated with a higher risk of developing IBH (p?=?0.0013). To our knowledge this is the first time in any atopic dermatitis suffering species, including man, where the same risk allele has been identified in two distinct populations.     

Microrheology of human synovial fluid of arthritis patients studied by diffusing wave spectroscopy

The viscoelastic properties of synovial fluid (SF) are critical to its functions of lubrication and shock‐absorption of joints in human body; a change in the viscoelastic properties, even of only a few percents, is often concomitant with arthritis. In this work, the elastic modulus G ′(f) and the viscous modulus G ′′(f) of SF from patients suffering from three kinds of joint diseases, namely, osteoarthritis (OA), rheumatoid arthritis (RA), and gouty arthritis (GA), were determined as a function of frequency “f ” (in the low frequency range from f ～ 0.1 to 10 Hz) by Diffusing Wave Spectroscopy (DWS) and correlated with the white blood cell (WBC) count. A strong correlation was observed, showing a higher WBC count corresponding to lower elastic and viscous moduli, G ′ and G ′′; further details depend on inflammatory vs. non‐inflammatory, and on the severity of inflammation. Different types of arthritis lead to different degrees of decreasing viscoelasticity. Identical measurements were carried out with a commercial visco‐supplementation (or artificial SF) to serve as reference. In general, the reduction in both G ′ and G ′′ was most severe in the case of GA and least severe in the case of OA. Besides, in all cases, the reduction in G ′ was more prominent than the reduction in G ′′, indicating that in general, the deterioration in the elasticity of SF by inflammation is more severe than that in the viscosity. This simple method for quantitative physical characterization of synovial fluid may serve as a useful complementary metric to the conventional biochemical analysis in clinical diagnosis of arthritis. (© 2012 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Evidence for a Rad18-independent frameshift mutagenesis pathway in human cell-free extracts

Bypass of replication blocks by specialized DNA polymerases is crucial for cell survival but may promote mutagenesis and genome instability. To gain insight into mutagenic sub-pathways that coexist in mammalian cells, we examined N-2-acetylaminofluorene (AAF)-induced frameshift mutagenesis by means of SV40-based shuttle vectors containing a single adduct. We found that in mammalian cells, as previously observed in E. coli, modification of the third guanine of two target sequences, 5'-GGG-3' (3G) and 5'-GGCGCC-3' (NarI site), induces -1 and -2 frameshift mutations, respectively. Using an in vitro assay for translesion synthesis, we investigated the biochemical control of these events. We showed that Pol eta, but neither Pol iota nor Pol zeta, plays a major role in the frameshift bypass of the AAF adduct located in the 3G sequence. By complementing PCNA-depleted extracts with either a wild-type or a non-ubiquitinatable form of PCNA, we found that this Pol eta-mediated pathway requires Rad18 and ubiquitination of PCNA. In contrast, when the AAF adduct is located within the NarI site, TLS is only partially dependent upon Pol eta and Rad18, unravelling the existence of alternative pathways that concurrently bypass this lesion.     

Diabetes with hypertension as risk factors for adult dengue hemorrhagic fever in a predominantly dengue serotype 2 epidemic: a case control study

Background

Dengue hemorrhagic fever (DHF) is a severe form of dengue, characterized by bleeding and plasma leakage. A number of DHF risk factors had been suggested. However, these risk factors may not be generalized to all populations and epidemics for screening and clinical management of patients at risk of developing DHF. This study explored demographic and comorbidity risk factors for DHF in adult dengue epidemics in Singapore in year 2006 (predominantly serotype 1) and in year 2007–2008 (predominantly serotype 2).

Methods

A retrospective case-control study was conducted with 149 DHF and 326 dengue fever (DF) patients from year 2006, and 669 DHF and 1,141 DF patients from year 2007–2008. Demographic and reported comorbidity data were collected from patients previously. We performed multivariate logistic regression to assess the association between DHF and demographic and co-morbidities for year 2006 and year 2007–2008, respectively.

Results

Only Chinese (adjusted odds ratio [AOR] = 1.90; 95% confidence interval [CI]: 1.01–3.56) was independently associated with DHF in year 2006. In contrast, age groups of 30–39 years (AOR = 1.41; 95% CI:1.09–1.81), 40–49 years (AOR = 1.34; 95% CI:1.09–1.81), female (AOR = 1.57; 95% CI:1.28–1.94), Chinese (AOR = 1.67; 95% CI:1.24–2.24), diabetes (AOR = 1.78; 95% CI:1.06–2.97), and diabetes with hypertension (AOR = 2.16; 95%CI:1.18–3.96) were independently associated with DHF in year 2007–2008. Hypertension was proposed to have effect modification on the risk of DHF outcome in dengue patients with diabetes. Chinese who had diabetes with hypertension had 2.1 (95% CI:1.07–4.12) times higher risk of DHF compared with Chinese who had no diabetes and no hypertension.

Conclusions

Adult dengue patients in Singapore who were 30–49 years, Chinese, female, had diabetes or diabetes with hypertension were at greater risk of developing DHF during epidemic of predominantly serotype 2. These risk factors can be used to guide triaging of patients who require closer clinical monitoring and early hospitalization in Singapore, when confirmed in more studies.     

Use of human perivascular stem cells for bone regeneration

Human perivascular stem cells (PSCs) can be isolated in sufficient numbers from multiple tissues for purposes of skeletal tissue engineering. PSCs are a FACS-sorted population of 'pericytes' (CD146+CD34-CD45-) and 'adventitial cells' (CD146-CD34+CD45-), each of which we have previously reported to have properties of mesenchymal stem cells. PSCs, like MSCs, are able to undergo osteogenic differentiation, as well as secrete pro-osteogenic cytokines. In the present protocol, we demonstrate the osteogenicity of PSCs in several animal models including a muscle pouch implantation in SCID (severe combined immunodeficient) mice, a SCID mouse calvarial defect and a femoral segmental defect (FSD) in athymic rats. The thigh muscle pouch model is used to assess ectopic bone formation. Calvarial defects are centered on the parietal bone and are standardly 4 mm in diameter (critically sized). FSDs are bicortical and are stabilized with a polyethylene bar and K-wires. The FSD described is also a critical size defect, which does not significantly heal on its own. In contrast, if stem cells or growth factors are added to the defect site, significant bone regeneration can be appreciated. The overall goal of PSC xenografting is to demonstrate the osteogenic capability of this cell type in both ectopic and orthotopic bone regeneration models.     

Considering water availability and the effect of solute concentration on high solids saccharification of lignocellulosic biomass

Milliliter scale (ligno)cellulose saccharifications suggest general solute concentration and its impact on water availability plays a significant role in detrimental effects associated with high solids lignocellulose conversions. A microtumbler developed to enable free‐fall mixing at dry solids loadings up to 35% (w/w) repeatedly produced known detrimental conversion trends on cellulose, xylan and pretreated lignocellulose with commercial enzymes. Despite this, high concentrations of insoluble nonhydrolysable dextrans did not depress saccharification extents in 5% (w/w) cellulose slurries suggesting mass transfer limitations may not significantly limit hydrolysis extents at high solids loadings. Interestingly, cellulose saccharification by purified cellulases showed increased conversions with increasing dry solids loadings. This prompted investigations into impacts the concentration of soluble species, such as sugar alcohols, low molecular weight enzyme preparation components, and monomer hydrolysis products, have on the hydrolysis environment. Such substances significantly depress conversion rates and were shown to correlatively lower water activity (A_w) in the hydrolysis environment while high insoluble solids concentrations did not. Furthermore, low‐field NMR on concentrated slurries of insoluble complex carbohydrates, including the nonhydrolysable dextrans, showed all solids constrained water significantly more than high concentrations of soluble species (inhibitory) suggesting water constraint may not be as problematic an issue at high solids loadings compared to the availability of water in the system. Additionally, the introduction of soluble species lessened overall water constraint in high solids systems and appears to shift the distribution of water away from insoluble surfaces. This is potentially a critical issue for industrial processes operating at high dry solids levels. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 2012     

Na,K-ATPase is a target of cigarette smoke and reduced expression predicts poor patient outcome of smokers with lung cancer

Diminished Na,K-ATPase expression has been reported in several carcinomas and has been linked to tumor progression. However, few studies have determined whether Na,K-ATPase function and expression are altered in lung malignancies. Because cigarette smoke (CS) is a major factor underlying lung carcinogenesis and progression, we investigated whether CS affects Na,K-ATPase activity and expression in lung cell lines. Cells exposed to CS in vitro showed a reduction of Na,K-ATPase activity. We detected the presence of reactive oxygen species (ROS) in cells exposed to CS before Na,K-ATPase inhibition, and neutralization of ROS restored Na,K-ATPase activity. We further determined whether Na,K-ATPase expression correlated with increasing grades of lung adenocarcinoma and survival of patients with smoking history. Immunohistochemical analysis of lung adenocarcinoma tissues revealed reduced Na,K-ATPase expression with increasing tumor grade. Using tissue microarray containing lung adenocarcinomas of patients with known smoking status, we found that high expression of Na,K-ATPase correlated with better survival. For the first time, these data demonstrate that CS is associated with loss of Na,K-ATPase function and expression in lung carcinogenesis, which might contribute to disease progression.