Novel understanding of ABC transporters ABCB1/MDR/P-glycoprotein, ABCC2/MRP2, and ABCG2/BCRP in colorectal pathophysiology

AIM: To evaluate ATP-binding cassette (ABC) transporters in colonic pathophysiology as they had recently been related to colorectal cancer (CRC) development. METHODS: Literature search was conducted on PubMed using combinations of the following terms: ABC transporters, ATP binding cassette transporter proteins, inflammatory bowel disease, ulcerative, colitis, Crohns disease, colorectal cancer, colitis, intestinal inflammation, intestinal carcinogenesis, ABCB1/P-glycoprotein (P-gp/CD243/MDR1), ABCC2/multidrug resistance protein 2 (MRP2) and ABCG2/breast cancer resistance protein (BCRP), Abcb1/Mdr1a, abcc2/Mrp2, abcg2/Bcrp, knock-out mice, tight junction, membrane lipid function. RESULTS: Recently, human studies reported that changes in the levels of ABC transporters were early events in the adenoma-carcinoma sequence leading to CRC. A link between ABCB1, high fat diet and gut microbes in relation to colitis was suggested by the animal studies. The finding that colitis was preceded by altered gut bacterial composition suggests that deletion of Abcb1 leads to fundamental changes of host-microbiota interaction. Also, high fat diet increases the frequency and severity of colitis in specific pathogen-free Abcb1 KO mice. The Abcb1 KO mice might thus serve as a model in which diet/environmental factors and microbes may be controlled and investigated in relation to intestinal inflammation. Potential molecular mechanisms include defective transport of inflammatory mediators and/or phospholipid translocation from one side to the other of the cell membrane lipid bilayer by ABC transporters affecting inflammatory response and/or function of tight junctions, phagocytosis and vesicle trafficking. Also, diet and microbes give rise to molecules which are potential substrates for the ABC transporters and which may additionally affect ABC transporter function through nuclear receptors and transcriptional regulation. Another critical role of ABCB1 was suggested by the finding that ABCB1 expression identifies a subpopulation of pro-inflammatory Th17 cells which were resistant to treatment with glucocorticoids. The evidence for the involvement of ABCC2 and ABCG2 in colonic pathophysiology was weak. CONCLUSION: ABCB1, diet, and gut microbes mutually interact in colonic inflammation, a well-known risk factor for CRC. Further insight may be translated into preventive and treatment strategies

<i>Cupriavidus pinatubonensis</i> AEO106 deals with copper-induced oxidative stress before engaging in biodegradation of the herbicide 4-chloro-2-methylphenoxyacetic acid

Abstract Background Microbial degradation of phenoxy acid (PA) herbicides in agricultural soils is important to minimize herbicide leaching to groundwater reservoirs. Degradation may, however, be hampered by exposure of the degrader bacteria to toxic metals as copper (Cu) in the soil environment. Exposure to Cu leads to accumulation of intracellular reactive oxygen species (ROS) in some bacteria, but it is not known how Cu-derived ROS and an ensuing oxidative stress affect the degradation of PA herbicides. Based on the previously proposed paradigm that bacteria deal with environmental stress before they engage in biodegradation, we studied how the degradation of the PA herbicide 2-methyl-4-chlorophenoxyacetic acid (MCPA) by the model PA degrader Cupriavidus pinatubonensis AEO106 was affected by Cu exposure. Results Exposure of C. pinatubonensis in batch culture to sublethal concentrations of Cu increased accumulation of ROS measured by the oxidant sensing probe 2,7-dichlorodihydrofluorescein diacetate and flow cytometry, and resulted in upregulation of a gene encoding a protein belong to the Ohr/OsmC protein family. The ohr/osmC gene was also highly induced by H2O2 exposure suggesting that it is involved in the oxidative stress response in C. pinatubonensis. The increased ROS accumulation and increased expression of the oxidative stress defense coincided with a delay in the catabolic performance, since both expression of the catabolic tfdA gene and MCPA mineralization were delayed compared to unexposed control cells. Conclusions The current study suggests that Cu-induced ROS accumulation in C. pinatubonensis activates a stress response involving the product of the ohr/osmC gene. Further, the stress response is launched before induction of the catabolic tfdA gene and mineralization occurs

eDNA Metabarcoding Benchmarked towards Conventional Survey Methods in Amphibian Monitoring

A keystone in protection work is accurate and thorough the monitoring of amphibian species, and the currently applied conventional survey methods are invasive, time-consuming, and dependent on expert knowledge. Research suggests that eDNA metabarcoding is a precise and cost-efficient method that could supplement the currently applied methods. The present study assessed the efficiency of conventional survey methods and eDNA metabarcoding in terms of species richness, the average number of detected species per site, the relative frequency of species occurrence, and the similarity of applied methods. The study found eDNA metabarcoding surveys to detect Lissotriton vulgaris (smooth newt), Triturus cristatus (great crested newt), Rana arvalis (moor frog), Rana temporaria (common frog), and Bufo bufo (common toad), as well as an average of 0.9 species per site, reflecting the species composition at the time of sampling in mid-July 2020. In addition to the species mentioned above, the conventional survey detected Epidalea calamita (natterjack toad) and an average of 1.7 species per site, reflecting the species composition at the time of sampling in early June 2020. The similarity between the methods applied in the present study was 27%, thus indicating a large number of unique observations of both eDNA metabarcoding and conventional surveys. The differences in detection can most likely be explained by the time of sampling, which was conducted a month apart. eDNA metabarcoding was efficient in detecting multiple amphibian species and produced unique observations that were not detected using conventional survey methods. Applying eDNA techniques as a supplement will most likely produce important knowledge on species distribution and presence, as well as enable more frequent monitoring due to cost efficiency and disturbance

eDNA Metabarcoding Benchmarked towards Conventional Survey Methods in Amphibian Monitoring

SIMPLE SUMMARY: Amphibian species are declining worldwide, and precise monitoring is key to ensuring timely protection and thereby ceasing deteriorating populations. Conventional monitoring methods are invasive, time-consuming, and dependent on expert knowledge. eDNA methods have been suggested as a replacement for or supplement to conventional survey methods. The present study assessed amphibian detection of conventional survey methods and eDNA metabarcoding in Danish lakes and ponds to address how the application of eDNA surveys can supplement the currently applied methodology. The study found eDNA metabarcoding to detect five out of six species detected through conventional methods. Furthermore, it is expected that the results in the present study reflect the time of sampling for the applied methods. The findings in the present study indicate that eDNA metabarcoding detects multiple Danish amphibian species and can produce knowledge on the occurrence and distribution for amphibian species. Implementing it as a supplement for conventional survey methods in nature monitoring will enable a higher frequency of monitoring and yield knowledge of species composition. ABSTRACT: A keystone in protection work is accurate and thorough the monitoring of amphibian species, and the currently applied conventional survey methods are invasive, time-consuming, and dependent on expert knowledge. Research suggests that eDNA metabarcoding is a precise and cost-efficient method that could supplement the currently applied methods. The present study assessed the efficiency of conventional survey methods and eDNA metabarcoding in terms of species richness, the average number of detected species per site, the relative frequency of species occurrence, and the similarity of applied methods. The study found eDNA metabarcoding surveys to detect Lissotriton vulgaris (smooth newt), Triturus cristatus (great crested newt), Rana arvalis (moor frog), Rana temporaria (common frog), and Bufo bufo (common toad), as well as an average of 0.9 species per site, reflecting the species composition at the time of sampling in mid-July 2020. In addition to the species mentioned above, the conventional survey detected Epidalea calamita (natterjack toad) and an average of 1.7 species per site, reflecting the species composition at the time of sampling in early June 2020. The similarity between the methods applied in the present study was 27%, thus indicating a large number of unique observations of both eDNA metabarcoding and conventional surveys. The differences in detection can most likely be explained by the time of sampling, which was conducted a month apart. eDNA metabarcoding was efficient in detecting multiple amphibian species and produced unique observations that were not detected using conventional survey methods. Applying eDNA techniques as a supplement will most likely produce important knowledge on species distribution and presence, as well as enable more frequent monitoring due to cost efficiency and disturbance

Neuronal substrates underlying stress resilience and susceptibility in rats

<div><p>Background</p><p>Stress and stressful life events have repeatedly been shown as causally related to depression. The Chronic Mild Stress rat model is a valid model of stress-induced depression. Like humans, rats display great heterogeneity in their response to stress and adversity. Hence some individuals are stress-sensitive and prone to develop depression-like behaviour in response to modest stressors, while others are stress-resilient and remain essentially symptom free.</p><p>Objectives</p><p>Compared to the large body of research, which describes stress-induced maladaptive neurobiological changes, relatively little attention has been devoted to understand resiliency to stress. The aim of the present study was to identify changes in neuronal activity, associated with stress-resilient and stress-susceptible chronic mild stress endophenotypes, by examining <i>c-Fos</i> expression in 13 different brain areas. Changes in <i>c-Fos</i> expression have been reported as associated to stressful conditions.</p><p>Methods</p><p>Stress-induced modulation of neuronal activation patterns in response to the chronic mild stress paradigm was mapped using the immediate early gene expression <i>c-Fos</i> as a marker. Quantification of the <i>c-Fos</i>-like immunoreactivity responses was done by semi-automated profile counting procedures and design-based stereology.</p><p>Results</p><p>Exposure to chronic mild stress significantly altered <i>c-Fos</i> expression in a total of 6 out of 13 investigated areas. Chronic mild stress was found to suppress the <i>c-Fos</i> response within the magnocellular ventral lateral geniculate nucleus of both stress subgroups. In the the lateral and ventral orbital cortices of stress-resilient rats, the <i>c-Fos</i> like immunoreactivity response was also repressed by stress exposure. On the contrary the <i>c-Fos</i> response within the amygdala, medial habenula, and infralimbic cortex was increased selectively for the stress-susceptible rats.</p><p>Conclusions</p><p>The study was initiated to characterize neuronal substrates associated with stress-coping mechanisms. Six areas, all of which represents limbic structures, were found to be sensitive to stress exposure. The effects within these areas associate to the hedonic status of the rats. Hence, these areas might be associated to stress-coping mechanisms underlying the chronic mild stress induced segregation into stress-susceptible and stress-resilient endophenotypes.</p></div

Figure modified from the BRAIN ATLAS [31].

<p>Pictures show 13 different substructures analysed at 3 rostrocaudal levels, within each structure, throughout different brain regions. In grey areas affected in stress-susceptible rats; in dark grey with stripes brain regions affected in both CMS phenotypes; in light grey areas affected in stress resilient rats. In beige, areas not affected by the CMS. DG, dendate gyrus; Cg1, cingulate cortex—area 1; IL, infralimbic cortex; PrL, prelimbic cortex; LO, lateral orbital cortex; VO, ventral orbital cortex; Pir, piriform cortex; PVA, paraventricular thalamic nucleus—anterior part; BLA, basolateral amygdaloid nucleus—anterior part; MHb, medial habenula; LHb, lateral habenula; CA3, field CA3 of hippocampus; VLGMC, ventral lateral geniculate nucleus magnocellular part; * p < 0.05; ** p < 0.01; *** p < 0.001.</p

Sucrose consumption following four weeks of the CMS regime.

<p>Four weeks exposure to chronic mild stressors resulted in a significant decrease of sucrose consumption of the anhedonic animals (n = 10) when compared to unchallenged controls (n = 10). The graph shows sucrose consumption of the control (n = 10), resilient (n = 10) and anhedonic-like (n = 10) animals at 4 different time points (week 1; week 2; week 3 and week 4). Significant differences on sucrose consumptions were detected among goups (p<0,0001) at different time points (p = 0,0084). The sucrose intake of anhedonic-like animals was significantly diminished when compared to control and resilient animals. Two-way ANOVA followed by post hoc Tukey´s multiple comparison test. * p<0,05; **p<0,01; ***p<0,001. Data is presented as mean (±SEM) sucrose intake, indexed to baseline values.</p

Representative microphotograph of c-Fos expression in different brain areas.

<p>High and low magnification pictures of representative sections to compare c-Fos-ir in eight brain regions. IL, infralimbic cortex; LO, lateral orbital cortex; VO, ventral orbital cortex; BLA, basolateral amygdaloid nucleus—anterior part; MHb, medial habenula; LHb, lateral habenula; CA3, field CA3 of hippocampus; VLGMC, ventral lateral geniculate nucleus; magnocellular part. Each panel shows a high and low magnification picture of the area, and a picture of the corresponding area modified from the rat brain atlas [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0179434#pone.0179434.ref031" target="_blank">31</a>]. Scale bar in panel A-H = 50um.</p

Study design showing the experimental outline and time course.

<p>(modified from [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0179434#pone.0179434.ref070" target="_blank">70</a>]).</p

Results summary.

<p>Results summary.</p