Inhibition of carnitine palmitoyl-transferase 1 is a potential target in a mouse model of Parkinson's disease.

Glucose metabolism is dysregulated in Parkinson's disease (PD) causing a shift toward the metabolism of lipids. Carnitine palmitoyl-transferase 1A (CPT1A) regulates the key step in the metabolism of long-chain fatty acids. The aim of this study is to evaluate the effect of downregulating CPT1, either genetically with a Cpt1a P479L mutation or medicinally on PD using chronic rotenone mouse models using C57Bl/6J and Park2 knockout mice. We show that Cpt1a P479L mutant mice are resistant to rotenone-induced PD, and that inhibition of CPT1 is capable of restoring neurological function, normal glucose metabolism, and alleviate markers of PD in the midbrain. Furthermore, we show that downregulation of lipid metabolism via CPT1 alleviates pathological motor and non-motor behavior, oxidative stress, and disrupted glucose homeostasis in Park2 knockout mice. Finally, we confirm that rotenone induces gut dysbiosis in C57Bl/6J and, for the first time, in Park2 knockout mice. We show that this dysbiosis is alleviated by the downregulation of the lipid metabolism via CPT1.

Targeted Next Generation Sequencing for Human Papillomavirus Genotyping in Cervical Liquid-Based Cytology Samples.

At present, human papillomavirus (HPV) testing is replacing morphology-based cytology as the primary tool for cervical cancer screening in several countries. However, the HPV assays approved for screening lack detection for all but one of the possibly carcinogenic HPV types and do not genotype all included HPV types. This study demonstrates the use of a targeted HPV next generation sequencing (NGS) panel to detect and genotype all 25 carcinogenic, probably carcinogenic, and possibly carcinogenic HPV types as well as the low-risk types HPV6 and HPV11. The panel was validated using a cohort of 93 paired liquid-based cytology samples (general practitioner (GP)-collected cervical samples and cervico-vaginal self-samples (SS)). Overall, the targeted panel had a sensitivity (GP = 97.7%, SS = 92.1%) and specificity (GP = 98.0%, SS = 96.4%) similar to the commercial HPV assays, Cobas® 4800 HPV DNA test (Roche) and CLART® HPV4S assay (GENOMICA). Interestingly, of the samples that tested positive with the NGS panel, three (6.4%) of the GP-collected samples and four (9.1%) of the self-samples tested positive exclusively for HPV types only included in the NGS panel. Thus, targeted HPV sequencing has great potential to improve the HPV screening programs since, as shown here, it can identify additional HPV positive cases, cases with HPV integration, variants in the HPV genome, and which HPV type is dominant in multi-infected cases.

Fabrication and characterization of extracellular matrix scaffolds obtained from adipose-derived stem cells.

Chronic non-healing wounds are detrimental for the quality of life of the affected individuals and represent a major burden for the health care systems. Adipose-derived stem cells (ASCs) are being investigated for the development of novel treatments of chronic wounds, as they have shown several positive effects on wound healing. While these effects appear to be mediated by the release of soluble factors, it is has also become apparent that the extracellular matrix (ECM) deposited by ASCs is essential in several phases of the wound healing process. In this work, we describe an approach to produce ECM scaffolds derived from ASCs in culture. Upon growth of ASCs into an overconfluent cell layer, a detergent-based cell extraction approach is applied to remove the cellular components. The extraction is followed by an enzymatic treatment to remove the residual DNA. The resultant cell-derived scaffolds are depleted of cellular components, display low DNA remnant, and retain the native fibrillar organization of the ECM. Analysis of the molecular composition of the ECM scaffolds revealed that they are composed of collagens type I and III, and fibronectin. The decellularized scaffolds represent a substrate that supports adhesion and proliferation of primary human fibroblasts and dermal microvascular endothelial cells, indicating their potential as platforms for wound healing studies.

Basolateral cholesterol depletion alters Aquaporin-2 post-translational modifications and disrupts apical plasma membrane targeting.

Apical plasma membrane accumulation of the water channel Aquaporin-2 (AQP2) in kidney collecting duct principal cells is critical for body water homeostasis. Posttranslational modification (PTM) of AQP2 is important for regulating AQP2 trafficking. The aim of this study was to determine the role of cholesterol in regulation of AQP2 PTM and in apical plasma membrane targeting of AQP2. Cholesterol depletion from the basolateral plasma membrane of a collecting duct cell line (mpkCCD14) using methyl-beta-cyclodextrin (MBCD) increased AQP2 ubiquitylation. Forskolin, cAMP or dDAVP-mediated AQP2 phosphorylation at Ser269 (pS269-AQP2) was prevented by cholesterol depletion from the basolateral membrane. None of these effects on pS269-AQP2 were observed when cholesterol was depleted from the apical side of cells, or when MBCD was applied subsequent to dDAVP stimulation. Basolateral, but not apical, MBCD application prevented cAMP-induced apical plasma membrane accumulation of AQP2. These studies indicate that manipulation of the cholesterol content of the basolateral plasma membrane interferes with AQP2 PTM and subsequently regulated apical plasma membrane targeting of AQP2.

Treatment decisions on Molar-Incisor Hypomineralization (MIH) by Norwegian dentists - a questionnaire study.

BACKGROUND: The study aimed to explore the variability between the treatment decisions dentists make for MIH-affected teeth. METHODS: In 2009, a pre-coded questionnaire was sent electronically to all dentists employed by the Public Dental Service (PDS) in Norway (n = 1061). The questions were related to treatment of MIH-affected teeth, including three patient cases illustrated by photographs and written case descriptions. RESULTS: Replies were obtained from 61.5 % of the respondents after two reminders. In the first case, showing a newly erupted first permanent molar with moderate hypomineralization and no disintegration of the surface enamel, the preferred treatment among the majority of the respondents (53.5 %) was application of fluoride varnish, while 19.6 % would seal the fissure with GIC material. In the second case, showing a severely damaged first permanent molar in a six year old child, more than half of the respondents (57.5 %) would place a conventional glass ionomer restoration and 10.5 % would use a stainless steel crown (SSC). In the third case, showing a severely damaged permanent first molar in a nine year old child, 43.8 % of the dentists would remove only the parts with soft, damaged enamel; while 35.2 % would remove more and 21.0 % would remove all affected enamel and leave the cavity margins in sound enamel. CONCLUSIONS: The survey shows that there is a wide disparity between clinicians' views on how MIH affected teeth should be treated. In a severely affected first permanent molar, only a minority of dentists would remove as much tooth substance as needed to get the full benefit of the acid etch pattern in sound enamel.

[Treatment of chronic constipation in children]. / Behandling af kronisk obstipation hos børn--en gennemgang af et Cochranereview.

Chronic functional constipation is a common disorder in childhood. The treatment most often consists of the modalities education, disimpaction and maintenance. In a recent Cochrane review of maintenance treatment polyethylene glycol (PEG) seems to be superior to placebo and to other laxatives including lactulose. This conclusion is in line with practice at most paediatric departments in Denmark. Still, larger randomized controlled studies with the curing of constipation as the primary end point are needed.

New insights into regulated aquaporin-2 function.

PURPOSE OF REVIEW: Aquaporin-2 (AQP2) water channels in principal cells of the kidney collecting duct are essential for urine concentration. Due to application of modern technologies, progress in our understanding of AQP2 has accelerated in recent years. In this article, we highlight some of the new insights into AQP2 function that have developed recently, with particular focus on the cell biological aspects of AQP2 regulation. RECENT FINDINGS: AQP2 is subjected to a number of regulated modifications, including phosphorylation and ubiquitination, which are important for AQP2 function, cellular localization and degradation. AQP2 is likely internalized via clathrin and non-clathrin-mediated endocytosis. Regulation of AQP2 endocytosis, in addition to exocytosis, is a vital mechanism in determining overall AQP2 membrane abundance. AQP2 is associated with regulated membrane microdomains. Studies using membrane cholesterol depleting reagents, for example statins, have supported the role of membrane rafts in regulation of AQP2 trafficking. Noncanonical roles for AQP2, for example in epithelial cell migration, are emerging. SUMMARY: AQP2 function and thus urine concentration is dependent on a variety of cell signalling mechanisms, posttranslational modification and interplay between AQP2 and its lipid environment. This complexity of regulation allows fine-tuning of AQP2 function and thus body water homeostasis.

Constipation in children with neurofibromatosis type 1.

BACKGROUND AND OBJECTIVES: Neurofibromatosis type 1 (NF1) is a hereditary, heterogenic, and multiorganic disease. The NF1 phenotype shows great variability in expressivity and often includes symptoms from the central and peripheral nervous systems. Bowel symptoms have been reported, but gastrointestinal function in NF1 remains to be described in detail. In this first systematic study of bowel function in children with NF1, we aimed to investigate symptoms of constipation and test the hypotheses that children with NF1 have abnormally large rectum and prolonged colonic transit time (CTT). METHODS: A total of 20 children with NF1 (age 8.2 ± 2.4 years) were evaluated with medical history; clinical examination; digital rectal examination; bowel and dietary diaries; Rome III criteria; measurement of rectal diameter by transabdominal ultrasound; and radiographic estimation of CTT. The control group for assessment of rectal diameter comprised 23 healthy children (mean age 9.1 ± 2.7 years). RESULTS: A total of 6 children with NF1 (30%) were constipated according to Rome III criteria. Average rectal diameter was significantly larger than for healthy children (32.9 ± 7.2 mm vs 21.4 ± 5.9 mm, P < 0.0001). Median CTT in NF1 children was 53 hours (range 26-101). Compared with existing normative data, CTT was prolonged (>84 hours) in 3 (19%). CONCLUSIONS: Symptoms of constipation were surprisingly common in children with NF1. Correspondingly, rectal diameters were abnormally large and a higher proportion than expected had prolonged CTT. The underlying pathophysiology remains obscure, but we hypothesise that abnormalities of the enteric nervous system or disturbed cellular growth could be present.