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Abstract: Objective: Ras proteins are known to affect cellular growth and function. The influence of the prenylation status of Ras on the observed changes in endothelial cell growth under high glucose conditions has not previously been examined. Methods: Human umbilical vein endothelial cells were exposed to normal or high glucose conditions for 72h. They were then examined for proliferative and hypertrophic effects, transforming growth factor β1 (TGFβ1) release, and phosphorylated p38 expression. The importance of prenylation was explored by the addition of mevalonate, isoprenoids or farnesyltransferase inhibitors to control the high glucose media and by measuring changes induced by high glucose and exogenous TGFβ1 in Ras prenylation and farnesyltransferase activity. Kidneys from diabetic rats treated with atorvastatin were also compared to specimens from untreated animals and the expression of the Ras effector p-Akt examined. Results: High glucose conditions caused a reduction in cell number. This was reversed in the presence of mevalonate or farnesylpyrophosphate (FPP), suggesting that the cell growth abnormalities observed are due to high glucose induced inhibition of the mevalonate pathway and subsequent prenylation of proteins. Endothelial cells exposed to high glucose increased their secretion of TGFβ1 and the phosphorylation of p38 both of which were reversed by concurrent exposure to FPP. A reduction in farnesyltransferase activity was observed after exposure to both high glucose and TGFβ1. Exposure to a farnesyltransferase inhibitor in control conditions mimicked the growth response observed with high glucose exposure and prenylated Ras was reduced by exposure to both high glucose and TGFβ1. Finally, interruption of the mevalonate pathway with a statin reduced the expression of p-Akt in diabetic rat kidneys. Conclusion: This study demonstrates that high glucose induced significant alterations in endothelial cell growth by inhibition of the mevalonate pathway, which subsequently mediates the increase in TGFβ1 and inhibition of Ras prenylation. [Copyright &y& Elsevier]

Our study compared methane (CH4) emissions from lactating dairy cows measured using the sulfur hexafluoride (SF6) tracer and open-circuit respiration chamber techniques. The study was conducted using 16 lactating Holstein-Friesian cows. In each chamber, the cow was fitted with the SF6 tracer apparatus to measure total CH4 emissions, including emissions from the rectum. Fresh ryegrass pasture was harvested daily and fed ad libitum to each cow with a supplement of 5 kg of grain/d. The CH4 emissions measured using the SF6 tracer technique were similar to those using the chamber technique: 331 vs. 322 g of CH4/d per cow. The accuracy of the SF6 tracer technique was indicated by considering the ratio of the CH4 emission measured using the SF6 tracer to the emission measured using the chamber for each cow on each day. The calculated ratio of 102.3% (SE = 1.51) was not different from 100%. A higher variability within cow between days was found for the SF6 tracer technique [coefficient of variation (CV) = 6.1%] than for the chamber technique (CV = 4.3%). The variability among cows was substantially higher than within cows, and was higher for the SF6 technique (CV = 19.6%) than for the chamber technique (CV = 17.8%). Our CH4 emission data were compared with whole-animal chamber studies conducted in Canada and Ireland. In the Canadian study the SF6 technique did not measure CH4 emissions from the rectum and emissions were 8% lower than those measured using the chamber, indicating that emissions from the rectum may be greater than previously measured (1%). The relationship between CH4 emission and dry matter intake was examined for our data and for that reported in the Canadian study. There was a difference in the slopes of the regressions derived from our data and that from Canada; 17.1 vs. 20.8 g of CH4/kg of dry matter intake. A difference between the 2 locations was expected based on the difference in diet composition for these 2 studies. The SF6 tracer technique is reasonably accurate for inventory purposes and for evaluating the effects of mitigation strategies on CH4 emissions. [ABSTRACT FROM AUTHOR]

The objective of this study was to conduct a life-cycle assessment (LCA) of greenhouse gas (GHG) emissions from a typical nongrazing dairy production system in Eastern Canada. Additionally, as dairying generates both milk and meat, this study assessed several methods of allocating emissions between these coproducts. An LCA was carried out for a simulated farm based on a typical nongrazing dairy production system in Quebec. The LCA was conducted over 6 yr, the typical lifespan of dairy cows in this province. The assessment considered 65 female Holstein calves, of which 60 heifers survived to first calving at 27 mo of age. These animals were subsequently retained for an average of 2.75 lactations. Progeny were also included in the analysis, with bulls and heifers in excess of replacement requirements finished as grain-fed veal (270 kg) at 6.5 mo of age. All cattle were housed indoors and fed forages and grains produced on the same farm. Prefarm gate GHG emissions and removals were quantified using Holos, a whole-farm software model developed by Agriculture and Agri-Food Canada and based on the Intergovernmental Panel for Climate Change Tier 2 and 3 methodologies with modifications for Canadian conditions. The LCA yielded a GHG intensity of 0.92 kg of CO2 Eq/kg of fat- and protein-corrected milk yield. Methane (CH4) accounted for 56% of total emissions, with 86% originating from enteric fermentation. Nitrous oxide accounted for 40% of total GHG emissions. Lactating cows contributed 64% of total GHG emissions, whereas calves under 12 mo contributed 10% and veal calves only 3%. Allocation of GHG emissions between meat and milk were assessed as (1) 100% allocation to milk, (2) economics, (3) dairy versus veal animals, and (4) International Dairy Federation equation using feed energy demand for meat and milk production. Comparing emissions from dairy versus veal calves resulted in 97% of the emissions allocated to milk. The lowest allocation of emissions to milk (78%) was associated with the International Dairy Federation equation. This LCA showed that greatest reductions in GHG emissions would be achieved by applying mitigation strategies to reduce enteric CH4 from the lactating cow, with minimal reductions being achievable in young stock. Choice of coproduct allocation method can also significantly affect the relative allocation of GHG emissions to milk and meat. [ABSTRACT FROM AUTHOR]

An experiment was conducted in vitro to determine whether the addition of saponin-containing Yucca schidigera or Quillaja saponaria reduces methane production without impairing ruminal fermentation or fiber digestion. A slightly lower dose of saponin was then fed to lactating dairy cows to evaluate effects on ruminal fermentation, methane production, total-tract nutrient digestibility, and milk production and composition. A 24-h batch culture in vitro incubation was conducted in a completely randomized design with a control (no additive, CON) and 3 doses of either saponin source [15, 30, and 45 g/kg of substrate dry matter (DM)] using buffered ruminal fluid from 3 dairy cows. The in vivo study was conducted as a crossover design with 2 groups of cows, 3 treatments, and three 28-d periods. Six ruminally cannulated cows were used in group 1 and 6 intact cows in group 2 (627 ± 55 kg of body weight and 155 ± 28 d in milk). The treatments were 1) early lactation total mixed ration, no additive (control; CON); 2) CON diet supplemented with whole-plant Y. schidigera powder at 10 g/kg of DM (YS); and 3) CON diet supplemented with whole-plant Q. saponaria powder at 10 g/kg of DM (QS). Methane production was measured in environmental chambers and with the sulfur hexafluoride (SF6) tracer technique. In vitro, increasing levels of both saponin sources decreased methane concentration in the headspace and increased the proportion of propionate in the buffered rumen fluid. Concentration of ammonia-N, acetate proportion, and the acetate:propionate ratio in the buffered rumen fluid as well as 24-h digestible neutral detergent fiber were reduced compared with the CON treatment. Medium and high saponin levels decreased DM digestibility compared with the CON treatment. A lower feeding rate of both saponin sources (10 g/kg of DM) was used in vivo in an attempt to avoid potentially negative effects of higher saponin levels on feed digestibility. Feeding saponin did not affect milk production, total-tract nutrient digestibility, rumen fermentation, or methane production. However, DM intake was greater for cows fed YS and QS than for CON cows, with a tendency for greater DM intake for cows fed YS compared with those fed QS. Consequently, efficiency of milk production (kg of milk/kg of DM intake) was lower for cows fed saponin compared with controls. The results show that although saponin from Y. schidigera and Q. saponaria lowered methane production in vitro, the reduction was largely due to reduced ruminal fermentation and feed digestion. Feeding a lower dose of saponin to lactating dairy cows avoided potentially negative effects on ruminal fermentation and feed digestion, but methane production was not reduced. Lower efficiency of milk production of cows fed saponin, and potential reductions in feed digestion at high supplementation rates may make saponin supplements an unattractive option for lowering methane production in vivo. [ABSTRACT FROM AUTHOR]

We examined the effects of monensin, provided by controlled-release capsules, on the enteric methane emissions and milk production of dairy cows receiving ryegrass pasture and grain. In a grazing experiment, 60 Holstein-Friesian cows were assigned randomly to 1 of 2 groups (control or monensin). Cows in the monensin group received 2 controlled-release capsules, with the second capsule administered 130 d after the first. Milk production was measured for 100 d following insertion of each capsule. The sulfur hexafluoride tracer gas technique was used to measure enteric methane emissions for 4 d starting on d 25 and 81 after insertion of the first capsule, and on d 83 after insertion of the second capsule. All cows grazed together as a single herd on a predominantly ryegrass sward and received 5 kg/d of grain (as-fed basis). In a second experiment, 7 pairs of lactating dairy cows (control and monensin) were used to determine the effects of monensin controlled-release capsules on methane emissions and dry matter intake. Methane emissions were measured on d 75 after capsule insertion by placing cows in respiration chambers for 3 d. Cows received fresh ryegrass pasture harvested daily and 5 kg/d of grain. The release rate of monensin from the capsules used in both experiments was 240 ± 0.072 mg/d, determined over a 100-d period in ruminally cannulated cows. The monensin dose was calculated to be 12 to 14.5 mg/kg of dry matter intake. There was no effect of monensin on methane production in either the grazing experiment (g/d, g/kg of milk solids) or the chamber experiment (g/d, g/kg of dry matter intake). In the grazing study, there was no effect of monensin on milk yield, but monensin increased milk fat yield by 51.5 g/d and tended to increase milk protein yield by 18.5 g/d. Monensin controlled-release capsules improved the efficiency of milk production of grazing dairy cows by increasing the yield of milk solids. However, a higher dose rate of monensin may be needed to reduce methane emissions from cows grazing pasture. [ABSTRACT FROM AUTHOR]

Severe lupus pneumonitis is a rare and life-threatening complication of systemic lupus erythematosus (SLE), characterized by its rapid progression and high mortality rate. This case report describes the clinical trajectory and therapeutic management of a young Aboriginal female with established lupus nephritis who developed severe lupus pneumonitis. Despite her stable renal condition under long-term immunosuppressive treatment, she experienced acute respiratory distress, leading to her admission to the intensive care unit and subsequent mechanical ventilation. The diagnostic process was complicated by the difficulty in obtaining tissue biopsies, necessitating reliance on clinical judgement and radiological evidence to formulate a diagnosis. The patient was treated with pulsed intravenous methylprednisolone followed by rituximab infusions, resulting in significant clinical and radiological improvement. This case highlights the importance of early and intensive immunosuppressive therapy in the management of severe lupus pneumonitis and underscores the utility of a multidisciplinary approach in overcoming diagnostic ambiguities. Furthermore, it contributes to the growing body of evidence supporting the efficacy of rituximab in severe lupus pneumonitis cases, offering insights into potential therapeutic avenues when conventional management strategies are inadequate or unsuitable., Competing Interests: None declared., (© 2024 The Author(s). Respirology Case Reports published by John Wiley & Sons Australia, Ltd on behalf of The Asian Pacific Society of Respirology.)

Melanoma transmitted through organ transplantation is an increasingly reported event. Immunosuppression increases the risk of melanoma; however, transmission of malignancy from transplanted organs is a distinct etiology of melanoma occurrence. The risk of transmission of melanoma from an organ donor with melanoma has yet to be determined. The authors aimed to investigate this phenomenon by reviewing the outcomes of patients that received organs from donors with melanoma. A systematic literature review was conducted with emphasis on identifying organ donors with known histories of melanoma and reported information regarding recipients of their organs. The databases PubMed, MEDLINE, Embase, and JBI EBP were searched in January 2023. Search terms included "melanoma," terms for solid organs, "donor," "transplant," "transmission," and their variations as well as terms related to temporal relations. Inclusion criteria were articles that stated outcomes in organ recipients from donors that had a diagnosis of melanoma either pretransplant or postmortem. Reference lists of selected articles were hand searched for further studies. A total of 232 articles were identified from the search parameters. After applying inclusion and exclusion criteria, 13 articles were selected. Hand searching the references of these articles yielded four additional articles. Of the 75 organ recipients that received organs from donors with known melanoma, 43 developed melanoma. While a definitive quantitative risk cannot be ascertained based on our review, the numerous reported cases of melanoma in organ recipients from donors that have melanoma should still be considered by clinicians., (© 2023 the International Society of Dermatology.)

Background: The gold standard of commencing hemodialysis with a functional arteriovenous fistula (AVF) is challenging. We aim to review factors associated with functional AVF at hemodialysis start at a tertiary hospital., Methods: We retrospectively reviewed incident hemodialysis patients or who had AVF creation at a single tertiary hospital from 2011 to 2016. Data was extracted for patient comorbidities, duration from referral to AVF creation and hemodialysis start, estimated glomerular filtration rate (eGFR) at surgical referral, referring nephrologist, events accelerating eGFR decline, and revisions for "failing to mature" AVF to assess factors associated with non-functioning AVF or late AVF creation, using multinomial logistic regression., Results: Two hundred two patients received hemodialysis and 51 had AVF creation but did not dialyze (AVF futility rate 20%). Of these, 133 (66%) commenced hemodialysis with a central venous catheter (CVC) and 69 (34%) with an AVF. Patients with functional AVFs at hemodialysis start were referred earlier than those with non-functional AVFs (median 256 vs 66 days before hemodialysis start, p  = 0.001). Age, sex, eGFR at surgical referral, and comorbidities were not predictive of patients with functional AVFs. Events accelerating eGFR decline were associated with an increased incidence of CVC at hemodialysis start (risk ratio (RR) 4.21, 95% confidence interval (CI) 1.96-9.03, p  < 0.0001). Referring nephrologists external to our renal unit may be associated with non-functional AVF at hemodialysis start (RR 6.60, 95% CI 1.74-25.13, p  = 0.006)., Conclusions: We found that functional AVFs required referral a median of 256 days prior to hemodialysis start and events accelerating eGFR decline increase the incidence of CVC at hemodialysis start. Age, sex, eGFR at surgical referral, and comorbidities did not inform the likelihood of timely AVF creation and evaluation of further predictive pre-dialysis factors is necessary to identify patients requiring early AVF creation whilst minimizing the cost of unnecessary procedures.

Aims: Our objective was to better understand the genetic bases of dilated cardiomyopathy (DCM), a leading cause of systolic heart failure., Methods and Results: We conducted the largest genome-wide association study performed so far in DCM, with 2719 cases and 4440 controls in the discovery population. We identified and replicated two new DCM-associated loci on chromosome 3p25.1 [lead single-nucleotide polymorphism (SNP) rs62232870, P = 8.7 × 10-11 and 7.7 × 10-4 in the discovery and replication steps, respectively] and chromosome 22q11.23 (lead SNP rs7284877, P = 3.3 × 10-8 and 1.4 × 10-3 in the discovery and replication steps, respectively), while confirming two previously identified DCM loci on chromosomes 10 and 1, BAG3 and HSPB7. A genetic risk score constructed from the number of risk alleles at these four DCM loci revealed a 3-fold increased risk of DCM for individuals with 8 risk alleles compared to individuals with 5 risk alleles (median of the referral population). In silico annotation and functional 4C-sequencing analyses on iPSC-derived cardiomyocytes identify SLC6A6 as the most likely DCM gene at the 3p25.1 locus. This gene encodes a taurine transporter whose involvement in myocardial dysfunction and DCM is supported by numerous observations in humans and animals. At the 22q11.23 locus, in silico and data mining annotations, and to a lesser extent functional analysis, strongly suggest SMARCB1 as the candidate culprit gene., Conclusion: This study provides a better understanding of the genetic architecture of DCM and sheds light on novel biological pathways underlying heart failure., (Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2021. For permissions, please email: journals.permissions@oup.com.)

The ubiquitin-proteasome system degrades ubiquitin-modified proteins to maintain protein homeostasis and to control signalling. Whole-genome sequencing of patients with severe deafness and early-onset cataracts as part of a neurological, sensorial and cutaneous novel syndrome identified a unique deep intronic homozygous variant in the PSMC3 gene, encoding the proteasome ATPase subunit Rpt5, which lead to the transcription of a cryptic exon. The proteasome content and activity in patient's fibroblasts was however unaffected. Nevertheless, patient's cells exhibited impaired protein homeostasis characterized by accumulation of ubiquitinated proteins suggesting severe proteotoxic stress. Indeed, the TCF11/Nrf1 transcriptional pathway allowing proteasome recovery after proteasome inhibition is permanently activated in the patient's fibroblasts. Upon chemical proteasome inhibition, this pathway was however impaired in patient's cells, which were unable to compensate for proteotoxic stress although a higher proteasome content and activity. Zebrafish modelling for knockout in PSMC3 remarkably reproduced the human phenotype with inner ear development anomalies as well as cataracts, suggesting that Rpt5 plays a major role in inner ear, lens and central nervous system development., (© 2020 The Authors. Published under the terms of the CC BY 4.0 license.)

Liquid manure is a significant source of methane (CH 4 ), a greenhouse gas. Many livestock farms use manure additives for practical and agronomic purposes, but the effect on CH 4 emissions is unknown. To address this gap, two lab studies were conducted, evaluating the CH 4 produced from liquid dairy manure with Penergetic-g ® (12 mg/L, 42 mg/L, and 420 mg/L) or AgrimestMix ® (30.3 mL/L). In the first study, cellulose produced 378 mL CH 4 /g volatile solids (VS) at 38 °C and there was no significant difference with Penergetic-g ® at 12 mg/L or 42 mg/L. At the same temperature, dairy manure produced 254 mL CH 4 /g VS and was not significantly different from 42 mg/L Penergetic-g ® . In the second lab study, the dairy manure control produced 187 mL CH 4 /g VS at 37 °C and 164 mL CH 4 /g VS at 20 °C, and there was no significant difference with AgrimestMix (30.3 mL/L) or Penergetic-g ® (420 mg/L) at either temperature. Comparisons of manure composition before and after incubation indicated that the additives had no effect on pH or VS, and small and inconsistent effects on other constituents. Overall, neither additive affected CH 4 production in the lab. The results suggest that farms using these additives are likely to have normal CH 4 emissions from stored manure.

Purpose: The purpose of this systematic review is to identify quality-of-life issues that affect participation in age-appropriate activities in chronically ill children, as reported by the children and their families., Summary of Key Points: Social and emotional functioning scores on the Pediatric Quality of Life Inventory (PedsQL) 4.0 were found to have the greatest frequency of poor agreement between parents and children in 4 of the 6 studies included in this review, suggesting parents and children have wide variation in their assessment in these areas of psychosocial function., Conclusions: Cumulative evidence appears to indicate that parents of children with chronic illness perceive their children as having a poorer quality of life than the children report for themselves., Recommendations for Clinical Practice: Identifying differences and commonalities between these reports can guide health care practitioners to specific activities that should be the focus of caring for children; specifically, functional goal development can become more personalized and appropriate.

Ciliopathies represent a wide spectrum of rare diseases with overlapping phenotypes and a high genetic heterogeneity. Among those, IFT140 is implicated in a variety of phenotypes ranging from isolated retinis pigmentosa to more syndromic cases. Using whole-genome sequencing in patients with uncharacterized ciliopathies, we identified a novel recurrent tandem duplication of exon 27-30 (6.7 kb) in IFT140, c.3454-488&#95;4182+2588dup p.(Tyr1152&#95;Thr1394dup), missed by whole-exome sequencing. Pathogenicity of the mutation was assessed on the patients' skin fibroblasts. Several hundreds of patients with a ciliopathy phenotype were screened and biallelic mutations were identified in 11 families representing 12 pathogenic variants of which seven are novel. Among those unrelated families especially with a Mainzer-Saldino syndrome, eight carried the same tandem duplication (two at the homozygous state and six at the heterozygous state). In conclusion, we demonstrated the implication of structural variations in IFT140-related diseases expanding its mutation spectrum. We also provide evidences for a unique genomic event mediated by an Alu-Alu recombination occurring on a shared haplotype. We confirm that whole-genome sequencing can be instrumental in the ability to detect structural variants for genomic disorders., (© 2018 Wiley Periodicals, Inc.)

Background: Preoperative anaemia is a risk factor for poorer postoperative outcomes and many colorectal cancer patients have iron-deficiency anaemia. The aim of this study was to assess if a preoperative iron-deficiency anaemia management protocol for elective colorectal surgery patients helps improve detection and treatment of iron-deficiency, and improve patient outcomes., Materials and Methods: Retrospective data was collected from 95 consecutive patients undergoing colorectal cancer surgery to establish baseline anaemia correction rates and perioperative transfusion rates. A new pathway for early detection of iron-deficiency anaemia, and treatment with intravenous iron replacement, for colorectal cancer patients was then developed and implemented. Data from 81 patients was collected prospectively post-implementation to assess the impact of the pathway., Results: Pre-intervention data showed anaemic patients were seventeen times more likely to require perioperative transfusion than non-anaemic patients (95% CI 1.9-151.0, p = 0.011). Post-intervention, fifteen patients with iron-deficiency were treated with either intravenous (n = 8) or oral iron (n = 7). Mean Day 3 postoperative haemoglobin levels were significantly lower in patients with uncorrected anaemia (9.5 g/dL, p = 0.004); those patients whose anaemia was corrected by iron replacement therapy preoperatively had similar postoperative results to non-anaemic patients (10.93 g/dL vs 11.4 g/dL, p = 0.781). Postoperative transfusion rates remained high at 38% in patients with uncorrected anaemia, compared to 0% in corrected anaemia and 3.5% in non-anaemic patients., Conclusions: Introduction of an iron-deficiency anaemia management pathway has resulted in improved perioperative haemoglobin levels, with a reduction in perioperative transfusion, in elective colorectal patients. Implementation of this pathway could result in similar outcomes across other categories of surgical patients., (Copyright © 2016 IJS Publishing Group Ltd. Published by Elsevier Ltd. All rights reserved.)

Background: Low bone mineral density (BMD) is a known independent predictor of mortality in the general elderly population. However, studies in patients with end-stage renal disease (ESRD) are limited. The present study evaluated mortality during long-term follow-up in a population of patients having dialysis for ESRD, in whom BMD was also measured., Methods: Fifty-eight patients with ESRD were recruited consecutively from a dialysis clinic and followed prospectively for 6 years. Baseline BMD of the lumbar spine and femoral neck (FN) were measured by X-ray absorptiometry and by peripheral quantitative CT at the radius and tibia. Serum calcium, phosphate, parathyroid hormone (PTH), and albumin were measured at baseline., Results: During follow-up, 25 patients died. Univariate analysis showed that mortality was significantly associated with FN-BMD: hazards ratio (HR) per 0.1 g/cm2 decrease 1.50 (95% CI 1.07-2.10), p = 0.019; FN-T score: HR per 1-SD decrease 1.84 (95% CI 1.16-2.92), p = 0.009; and tibial cortical density: HR per 10 mg/cm3 decrease 1.08 (95% CI 1.02-1.14), p = 0.010. In multivariate analysis with stepwise adjustment for age, sex, transplant status, albumin, PTH, phosphate, dialysis duration, diabetes, and smoking, FN-T score remained significantly associated with mortality: HR per 1-SD decrease 1.82 (95% CI 1.02-3.24), p = 0.044, whereas the HR for FN-BMD and tibial cortical density were no longer significant. When 4 patients who had peritoneal dialysis were excluded, the HR relating FN-BMD, FN-T score, and tibial cortical density to mortality remained significant but became insignificant when albumin was included in the multivariate analysis., Conclusion: Reduced FN-BMD, FN-T score, and tibial cortical density were significantly associated with an increased risk of death in patients with ESRD., (© 2017 S. Karger AG, Basel.)

The REvolutionary Approaches and Devices for Nucleic Acid analysis (READNA) project received funding from the European Commission for 41/2 years. The objectives of the project revolved around technological developments in nucleic acid analysis. The project partners have discovered, created and developed a huge body of insights into nucleic acid analysis, ranging from improvements and implementation of current technologies to the most promising sequencing technologies that constitute a 3(rd) and 4(th) generation of sequencing methods with nanopores and in situ sequencing, respectively., (Copyright © 2015. Published by Elsevier B.V.)

For decades, ill-defined autosomal dominant renal diseases have been reported, which originate from tubular cells and lead to tubular atrophy and interstitial fibrosis. These diseases are clinically indistinguishable, but caused by mutations in at least four different genes: UMOD, HNF1B, REN, and, as recently described, MUC1. Affected family members show renal fibrosis in the biopsy and gradually declining renal function, with renal failure usually occurring between the third and sixth decade of life. Here we describe 10 families and define eligibility criteria to consider this type of inherited disease, as well as propose a practicable approach for diagnosis. In contrast to what the frequently used term 'Medullary Cystic Kidney Disease' implies, development of (medullary) cysts is neither an early nor a typical feature, as determined by MRI. In addition to Sanger and gene panel sequencing of the four genes, we established SNaPshot minisequencing for the predescribed cytosine duplication within a distinct repeat region of MUC1 causing a frameshift. A mutation was found in 7 of 9 families (3 in UMOD and 4 in MUC1), with one indeterminate (UMOD p.T62P). On the basis of clinical and pathological characteristics we propose the term 'Autosomal Dominant Tubulointerstitial Kidney Disease' as an improved terminology. This should enhance recognition and correct diagnosis of affected individuals, facilitate genetic counseling, and stimulate research into the underlying pathophysiology.

Background: Detection of B cell clonality is useful for assisting in the diagnosis of B cell lymphomas. Clonality assessment can be accomplished through evaluation of KAPPA and LAMBDA light chain expression. Currently, only slide based methods are available for the majority of patient biopsies and do not detect light chain protein or mRNA in many B-cell lymphomas. Herein we evaluated a new method, known as colorimetric in situ hybridization (CISH), with improved sensitivity and multiplexing capacity, for its usefulness in clonality detection in mature B cell malignancies., Methods: The KAPPA and LAMBDA ISH was performed on a Ventana Benchmark XT utilizing two color chromogenetic detection. The probes comprised 2 haptenated riboprobes each approximately 500 base pairs long directed against the conserved regions of either KAPPA or LAMBDA mRNA. The dual colors consisted of silver deposition (black) for KAPPA light chain and a novel (pink) chromogen for LAMBDA light chain. Following optimization, CISH allowed visualization of mRNA in benign B cells in reactive tissues including germinal center, mantle zone, and post-germinal center cells. We then identified 79 cases of B cell lymphoma with formalin-fixed paraffin-embedded (FFPE) biopsies including: follicular (36 cases), mantle cell (6 cases), marginal zone (12 cases), lymphoplasmacytic (6 cases), small lymphocytic (4 cases), and diffuse large B cell (15 cases), which were selected on the basis of either prior flow cytometry or immunohistochemistry (IHC) results to serve as the predicate, "gold standard," comparator., Results: 39/79 (49.4%) cases were classified as KAPPA and 29/79 (36.7%) as LAMBDA light chain restricted; while 9/79 (11.3%) cases were classified as indeterminate. Of the 70 cases with KAPPA or LAMBDA light chain restricted CISH, 69/70 (98.6%) were concordant with the reference method, while 1/70 (1.4%) was discordant., Conclusions: Optimized CISH detected lower levels of mRNA than can be visualized with current slide based methods, making clonality assessment in FFPE biopsies possible for mature B cell neoplasms. In this preliminary study, CISH was highly accurate compared to flow cytometry or IHC. CISH offers the possibility of wider applicability of light chain ISH and is likely to become a useful diagnostic tool., Virtual Slides: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1430491067123856.

Micrometeorological techniques can be applied to estimate methane (CH4) emissions from ruminants and livestock manure using CH4 concentration measured within the internal surface boundary layer. The main advantage of these techniques is that they are non-intrusive, thereby eliminating the impact of the measurement set-up on the calculated CH4 emission. This review focuses on four micrometeorological techniques, namely, the integrated horizontal flux (IHF), flux gradient (FG), eddy covariance (EC) and the dispersion modelling using the backward Lagrangian stochastic method (BLS). Each technique has unique advantages and limitations when used for estimating enteric (ruminant) and manure CH4 emissions. The IHF technique may be theoretically simpler then the FG, EC or BLS techniques, but all require high-resolution instruments to measure concentration. The EC and BLS techniques also require a measurement of the wind statistics. This review discusses the appropriate use of these four micrometeorological techniques for estimating CH4 emissions in animal agriculture and the recent advances in measurement technology.

Nucleic acid sequencing is the mainstay of biological research. There are several generations of DNA sequencing technologies that can be well characterized through their nature and the kind of output they provide. Dideoxy terminator sequencing developed by Sanger dominated for 30 years and was the workhorse used for the Human Genome Project. In 2005 the first 2nd generation sequencer was presented with an output orders of magnitude higher than Sanger sequencing and dramatically decreased cost. We are now at the dawn of 3rd generation with nanopore systems that are being developed for DNA sequencing. Meanwhile the field is also broadening into applications that complement 1st, 2nd and 3rd generation sequencing systems to get high resolution genetic information. The REvolutionary Approaches and Devices for Nucleic Acid analysis (READNA) consortium funded by the European Commission under FP7 has made great contributions to the development of new nucleic acid analysis methodology., (Copyright © 2012 Elsevier B.V. All rights reserved.)

The objective of this study was to conduct a life-cycle assessment (LCA) of greenhouse gas (GHG) emissions from a typical nongrazing dairy production system in Eastern Canada. Additionally, as dairying generates both milk and meat, this study assessed several methods of allocating emissions between these coproducts. An LCA was carried out for a simulated farm based on a typical nongrazing dairy production system in Quebec. The LCA was conducted over 6 yr, the typical lifespan of dairy cows in this province. The assessment considered 65 female Holstein calves, of which 60 heifers survived to first calving at 27 mo of age. These animals were subsequently retained for an average of 2.75 lactations. Progeny were also included in the analysis, with bulls and heifers in excess of replacement requirements finished as grain-fed veal (270 kg) at 6.5 mo of age. All cattle were housed indoors and fed forages and grains produced on the same farm. Pre-farm gate GHG emissions and removals were quantified using Holos, a whole-farm software model developed by Agriculture and Agri-Food Canada and based on the Intergovernmental Panel for Climate Change Tier 2 and 3methodologies with modifications for Canadian conditions. The LCA yielded a GHG intensity of 0.92 kg of CO(2) Eq/kg of fat- and protein-corrected milk yield. Methane (CH(4)) accounted for 56% of total emissions, with 86% originating from enteric fermentation. Nitrous oxide accounted for 40% of total GHG emissions. Lactating cows contributed 64% of total GHG emissions, whereas calves under 12 mo contributed 10% and veal calves only 3%. Allocation of GHG emissions between meat and milk were assessed as (1) 100% allocation to milk, (2) economics, (3) dairy versus veal animals, and (4) International Dairy Federation equation using feed energy demand for meat and milk production. Comparing emissions from dairy versus veal calves resulted in 97% of the emissions allocated to milk. The lowest allocation of emissions to milk (78%) was associated with the International Dairy Federation equation. This LCA showed that greatest reductions in GHG emissions would be achieved by applying mitigation strategies to reduce enteric CH(4) from the lactating cow, with minimal reductions being achievable in young stock. Choice of coproduct allocation method can also significantly affect the relative allocation of GHG emissions to milk and meat., (Copyright © 2012 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Interleukin 7 (IL7) is a critical factor for lymphocyte homeostasis. A dysfunction of the IL7/IL7R pathway has previously been described in HIV-1 infection, and promising results were observed in recent analyses of IL7 for therapeutic use in HIV infected individuals. However, further investigations are still warranted to understand the possible roles of this cytokine. Here, we explored whether the IL7 and IL7RA genetic polymorphisms were associated with the progression of HIV infection. We extensively genotyped the IL7 and IL7RA genes in the GRIV (Genomics of Resistance to Immunodeficiency Virus) cohort, composed of patients with extreme progression profiles - long-term non (LTNP) and rapid (RP) progressors--, and in a healthy control group (CTR). Statistical case-control analyses were performed using the Fisher's exact test, comparing either LTNP vs CTR or RP vs CTR. Three IL7RA SNPs (single nucleotide polymorphisms--rs7701176, rs987106 and rs10491434), but no IL7 SNPs, were significantly associated with rapid disease progression (P < 0.01). In a multi-marker analysis focusing on functional variants, a strong association between an IL7RA haplotype and rapid progression was observed (P = 5.59 x 10(-3)). In summary, our comprehensive genetic study revealed three SNPs and a risk of haplotype associated with rapid progression to AIDS in the IL7RA gene. Interestingly, the haplotype is composed of SNPs previously identified in other inflammatory diseases (e.g., multiple sclerosis) by GWAS and by functional studies. Our results contribute to the growing understanding of the role of IL7/IL7R in HIV disease progression, and more widely, in CD4+ T cell homeostasis.

The centrality of the transcriptional regulator Snail in epithelial-to-mesenchymal transformation (EMT), known to occur in models of diabetic nephropathy, has not been established. Transforming growth factor beta-1 (TGFbeta1) is induced in diabetic nephropathy and induces both Snail and EMT. Hypoxia inducible factors (HIFs) are known to induce Snail, independent of TGFbeta1. Notch induction is integral to Snail induction and EMT in tumour cells, but its role in the kidney is unknown. The present study was undertaken to determine the upstream regulators of Snail in the kidney in high glucose and hypoxic conditions. HK-2 cells were cultured in normoxic, hypoxic, high glucose and combined hypoxic/high glucose conditions. The expression of HIF1alpha, NotchIC, Snail, Lysyl oxidase-like 2 (Loxl2), and Hairy and Enhancer Split-1 (Hes1) were measured. We found that hypoxia increased HIF1alpha expression; however, concurrent exposure to high glucose blunted this effect. A similar pattern was observed in Lox12 expression, suggesting that Loxl2 was downstream of HIF1alpha, which was confirmed using siRNA techniques. Snail was upregulated by hypoxia and high glucose and in combination the effect was additive, suggesting independent upstream activation pathways by the two stimuli. Hes1 was upregulated by high glucose and to a lesser extent by hypoxia, but the effect of the combined stimuli was no greater than that observed with high glucose alone. NotchIC was downregulated by both hypoxia and high glucose, and in combination the effect was additive. Therefore, this study suggests that hypoxia and high glucose induce Snail expression through distinct pathways, independent of Notch signalling., (Crown Copyright 2010. Published by Elsevier Ltd. All rights reserved.)

Psoriasis is a common inflammatory and hyperproliferative skin disease. Recent studies have reported that common genetic factors may underlie both skin and immune-mediated disorders. We hypothesized that such genes may be involved in susceptibility to psoriasis, and undertook an association analysis of 22 candidate genes in a set of French high-risk psoriasis families. One hundred fifty-three single-nucleotide polymorphisms (SNPs) were genotyped and the transmission of alleles in nuclear families was analyzed using the FBAT (family-based association test). To further investigate suggestive associations, LNM (logistic-normal models) and MQLS (modified quasi-likelihood score) methods, which take the whole pedigree structure information of families into consideration, were also applied. Our study supported the involvement of six candidate genes in susceptibility to psoriasis: SCL12A8, which belongs to the solute carrier gene family; FLG and TGM5, which are involved in epidermal differentiation; CARD15 and CYLD, which modulate the transcription factor NF-kB; and IL1RN, which encodes an IL receptor antagonist. Furthermore, we found evidence for interaction between the major risk allele, HLA-Cw6, and CARD15, CYLD, and TGM5 susceptibility alleles. Taken together, our data show that shared genetic factors may contribute to the etiology of both psoriasis and other skin or immune-mediated disorders.

Chronic kidney disease is a major worldwide public health problem that causes substantial morbidity and mortality. Studies from the Asia-Pacific region have reported some of the highest chronic kidney disease prevalence rates in the world, but access to dialysis is limited in many countries, making it imperative to identify high-risk individuals. We performed a participant-level data overview of prospective studies conducted in the Asia-Pacific region to quantify the magnitude and direction of the associations between putative risk factors and renal death. Age- and sex-adjusted Cox proportional hazards models were applied to pooled data from 35 studies to calculate hazard ratios (95% CIs) for renal death associated with a standardized change in risk factors. Among 560 352 participants followed for a median of 6.8 years, a total of 420 renal deaths were observed. Continuous and positive associations among systolic blood pressure, diastolic blood pressure, fasting blood glucose, and total cholesterol levels with renal death were observed, as well as a continuous but inverse association with high-density lipoprotein cholesterol. Systolic blood pressure was the strongest risk factor for renal death with each SD increase in systolic blood pressure (19 mm Hg) associated with >80% higher risk (hazard ratio: 1.84; 95% CI: 1.60 to 2.12). Neither cigarette smoking nor excess weight was related to the risk of renal death (P>0.10). The results were similar for cohorts in Asia and Australia. These results suggest that primary prevention strategies for renal disease should focus on individuals with elevated blood pressure, diabetes mellitus, and dyslipidemia.

Objective: Genetic and environmental factors modulate the susceptibility to diabetic nephropathy, as initiating and/or progression factors. The objective of the European Rational Approach for the Genetics of Diabetic Complications (EURAGEDIC) study is to identify nephropathy susceptibility genes. We report molecular genetic studies for 127 candidate genes for nephropathy., Research Design and Methods: Polymorphisms were identified through sequencing of promoter, exon, and flanking intron gene regions and a database search. A total of 344 nonredundant SNPs and nonsynonymous variants were tested for association with diabetic nephropathy (persistent albuminuria >/=300 mg/24 h) in a large type 1 diabetes case/control (1,176/1,323) study from three European populations., Results: Only one SNP, rs2281999, located in the UNC13B gene, was significantly associated with nephropathy after correction for multiple testing. Analyses of 21 additional markers fully characterizing the haplotypic variability of the UNC13B gene showed consistent association of SNP rs13293564 (G/T) located in intron 1 of the gene with nephropathy in the three populations. The odds ratio (OR) for nephropathy associated with the TT genotype was 1.68 (95% CI 1.29-2.19) (P = 1.0 x 10(-4)). This association was replicated in an independent population of 412 case subjects and 614 control subjects (combined OR of 1.63 [95% CI 1.30-2.05], P = 2.3 x 10(-5))., Conclusions: We identified a polymorphism in the UNC13B gene associated with nephropathy. UNC13B mediates apopotosis in glomerular cells in the presence of hyperglycemia, an event occurring early in the development of nephropathy. We propose that this polymorphism could be a marker for the initiation of nephropathy. However, further studies are needed to clarify the role of UNC13B in nephropathy.

The adaptive immune response of ovarian cancer patients has been linked to survival, and is known to be impaired in the tumor microenvironment. Little is known about relationships between biobehavioral factors such as depressed mood and anxiety and the adaptive immune response in ovarian cancer. Thirty-seven patients with epithelial ovarian cancer and 14 patients with benign ovarian neoplasms completed psychosocial questionnaires pre-surgery. Lymphocytes from peripheral blood, tumor, and ascites (fluid around the tumor), were obtained on the day of surgery. Expression of the Type-1 cytokine interferon-gamma (IFN gamma), and the Type-2 cytokine interleukin-4 (IL-4) by T-helper (CD4(+)) and T-cytotoxic (CD8(+)) cells was measured under autologous tumor-stimulated, polyclonally-stimulated, or unstimulated conditions. Links with mood were examined. Among cancer patients, marked elevations in unstimulated and tumor-stimulated Type-2 responses were seen, particularly in ascites and tumor-infiltrating lymphocytes (P values<0.01). With polyclonal stimulation, lymphocytes from all compartments expressed elevated Type-1 cytokines (P values<0.014). Depressed and anxious mood were both associated with significantly lower ratios of polyclonally-stimulated CD4(+) cells producing IFN gamma (TH(1) cells) vs. IL-4 (TH(2) cells) in all compartments (depressed mood: P=0.012; anxiety: P=0.038) and depressed mood was also related to lower ratios of polyclonally-stimulated CD8(+) cells producing IFN gamma (TC(1)) vs. IL-4 (TC(2)) (P=0.035). Although effects of polyclonal stimulation should be generalized with caution to the in vivo immune response, findings suggest that depressed and anxious mood are associated with greater impairment of adaptive immunity in peripheral blood and in the tumor microenvironment among ovarian cancer patients.

Psychosocial factors are known to be associated with properties of both NK cells and T cells in cancer patients. Less is known about the relationship between psychosocial factors and NKT cells, a rare group of lymphocytes that have known relevance for tumor control. We examined four psychosocial factors and percentage and number of CD3+CD56+ NKT cells, CD3-CD56+ NK cells, and CD3+CD56- T cells in peripheral blood lymphocytes (PBL), ascites, and tumor of 35 ovarian cancer patients and 28 patients with benign pelvic masses. Patients awaiting surgery for a suspected cancerous mass completed questionnaires and gave a pre-surgical blood sample. Ascites and tumor were taken during surgery. After lymphocyte isolation, subpopulations were analyzed by flow cytometry. Benign and cancer patients did not differ on PBL subpopulations. Among cancer patients, NKT cell percentage was significantly higher in tumor and ascites than in PBL; T cell percentage was significantly higher in PBL than tumor. NKT, NK, and T cell number were significantly higher in peripheral blood than in ascites. Positive reframing was related to significantly higher NKT cell percentage and number in PBL. Social support was related to significantly higher NKT cell percentage in tumor. Vigor was related to significantly higher NKT cell percentage in PBL. Total mood disturbance was not related to NKT cell percentage or number. No significant relationships were found between psychosocial factors and NK cell percentage and number and T cell percentage and number. Given the anti-tumor activity of CD3+CD56+ cells, these relationships may have relevance for cancer control.

Our study compared methane (CH4) emissions from lactating dairy cows measured using the sulfur hexafluoride (SF6) tracer and open-circuit respiration chamber techniques. The study was conducted using 16 lactating Holstein-Friesian cows. In each chamber, the cow was fitted with the SF6 tracer apparatus to measure total CH4 emissions, including emissions from the rectum. Fresh ryegrass pasture was harvested daily and fed ad libitum to each cow with a supplement of 5 kg of grain/d. The CH4 emissions measured using the SF6 tracer technique were similar to those using the chamber technique: 331 vs. 322 g of CH4/d per cow. The accuracy of the SF6 tracer technique was indicated by considering the ratio of the CH4 emission measured using the SF6 tracer to the emission measured using the chamber for each cow on each day. The calculated ratio of 102.3% (SE = 1.51) was not different from 100%. A higher variability within cow between days was found for the SF6 tracer technique [coefficient of variation (CV) = 6.1%] than for the chamber technique (CV = 4.3%). The variability among cows was substantially higher than within cows, and was higher for the SF6 technique (CV = 19.6%) than for the chamber technique (CV = 17.8%). Our CH4 emission data were compared with whole-animal chamber studies conducted in Canada and Ireland. In the Canadian study the SF6 technique did not measure CH4 emissions from the rectum and emissions were 8% lower than those measured using the chamber, indicating that emissions from the rectum may be greater than previously measured (1%). The relationship between CH4 emission and dry matter intake was examined for our data and for that reported in the Canadian study. There was a difference in the slopes of the regressions derived from our data and that from Canada; 17.1 vs. 20.8 g of CH4/kg of dry matter intake. A difference between the 2 locations was expected based on the difference in diet composition for these 2 studies. The SF6 tracer technique is reasonably accurate for inventory purposes and for evaluating the effects of mitigation strategies on CH4 emissions.

Purpose: Psychosocial stress has been related to impaired immunity in cancer patients. However, the extent to which these relationships exist in immune cells in the tumor microenvironment in humans has not been explored. We examined relationships among distress, social support, and natural killer (NK) cell activity in ovarian cancer patients in peripheral-blood mononuclear cells (PBMC), ascitic fluid, and tumor-infiltrating lymphocytes (TIL)., Patients and Methods: Patients awaiting surgery for a pelvic mass suspected of being ovarian cancer completed psychological questionnaires and gave a presurgical sample of peripheral blood. Samples of tumor and ascites were taken during surgery, lymphocytes were then isolated, and NK cytotoxicity and percentage were determined. The final sample, which was confirmed by surgical diagnosis, included 42 patients with epithelial ovarian cancer and 23 patients with benign masses., Results: Peripheral NK cell activity was significantly lower among ovarian cancer patients than in patients with benign masses. Among ovarian cancer patients, NK cytotoxicity in TIL was significantly lower than in PBMC or ascitic fluid. Social support was related to higher NK cytotoxicity in PBMC and TIL, adjusting for stage. Distress was related to lower NK cytotoxicity in TIL. A multivariate model indicated independent associations of both distress and social support with NK cell activity in TIL., Conclusion: Psychosocial factors, such as social support and distress, are associated with changes in the cellular immune response, not only in peripheral blood, but also at the tumor level. These relationships were more robust in TIL. These findings support the presence of stress influences in the tumor microenvironment.

Background: Abnormalities in the structure and function of glomerular endothelial cells play a pivotal role in the development of progressive renal disease. The vascular abnormalities observed in the renal tubulointerstitium, however, correlate more strongly with progressive renal failure. Therefore, the successful isolation and culture of human renal microvascular endothelial cells from both the glomerulus and tubulointerstitium are paramount in studying renal disease models., Methods and Results: This study describes a simple and reproducible method for the isolation of human tubulointerstitial and glomerular endothelial cells by using immunomagnetic separation with anti-platelet endothelial-cell adhesion (anti-PECAM-1) Dyna beads, followed by manual weeding of mesangial and fibroblast contamination. No significant changes in morphological or immunohistochemical characteristics were observed up to passage two of culture. The in vitro characteristics of the endothelial cells were compared to the renal cortical endothelial cells in vivo and the standard human umbilical vein endothelial cell model (HUVECs). Similar to HUVECs, both populations of renal microvascular endothelial cells had a classical cobblestone appearance, stained positively for von Willebrand Factor and PECAM-1 and negatively for antifibroblast surface antigen and anticytokeratin. Differences in the expression of von Willebrand Factor, Wiebel Palade bodies and Flk-1 staining were observed between glomerular and tubulointerstitial endothelial cells. These immunohistochemical characteristics suggested that tubulointerstital endothelial cells were more closely aligned to HUVECS than to the glomerular endothelial cells. This observation indicated that HUVECs may be a suitable model for determining the tubulointerstitial endothelial response to systemic injury., Conclusion: In conclusion, a unique and novel method for the differential isolation of both glomerular and tubulointerstitial endothelial cells has been developed. Significantly, characterization of these populations suggests a role for HUVECS in the study of renal tubulointerstitial disease.

Human endothelial cells were exposed to 5 mM glucose (control), 25 mM (high) glucose, or osmotic control for 72 h. TGF-beta1 production, cell growth, death, and cell cycle progression, and the effects of TGF-beta1 and TGF-beta neutralization on these parameters were studied. High glucose and hyperosmolarity increased endothelial TGF-beta1 secretion (P < 0.0001) and bioactivity (P < 0.0001). However, high glucose had a greater effect on reducing endothelial cell number (P < 0.001) and increasing cellular protein content (P < 0.001) than the osmotic control. TGF-beta antibody only reversed the antiproliferative and hypertrophic effects of high glucose. High glucose altered cell cycle progression and cyclin-dependent kinase inhibitor expression independently of hyperosmolarity. High glucose increased endothelial cell apoptosis (P < 0.01), whereas hyperosmolarity induced endothelial cell necrosis (P < 0.001). TGF-beta antibody did not reverse the apoptotic effects observed with high glucose. Exogenous TGF-beta1 mimicked the increased S phase delay but not endoreduplication observed with high glucose. High glucose altered endothelial cell growth, apoptosis, and cell cycle progression. These growth effects occurred principally via a TGF-beta1 autocrine pathway. In contrast, apoptosis and endoreduplication occurred independently of this cytokine and hyperosmolarity.