Effect of Type of Pregnancy on Transcriptional and Plasma Metabolic Response in Sheep and Its Further Effect on Progeny Lambs.

The following study was performed in order to determine the effect of type of pregnancy on the transcriptional expression of genes that are engaged in angiogenesis and cell turnover/lactogenesis in the ewe mammary gland, evaluating its impact on the plasma metabolic response. In addition, an assessment of its further influence on plasma metabolic response, performance, and muscle transcriptional expression of lipogenic enzymes in progeny lambs was made. Thirteen Ile de France sheep (six twin- and seven single-bearing ewes) were allocated to graze ad libitum naturalized pasture from d 45 pre-partum to day 70 post-partum, while keeping their lambs on the same diet until day 60 after weaning. The samples were collected at different times and analyzed by qRT-PCR and plasma metabolic indicators. The data were processed using SPSS package. The results showed that twin-bearing ewes overexpressed VEGFR1 at birth, and BCL2 at birth and day 35 post-partum; however, single-bearing ewes overexpressed CAIV and IGF1 at day 35 post-partum. Similar metabolite concentrations in blood plasma were found between groups of ewes. The plasma metabolic response in lambs was similar between groups and it did not influence their performance, where a similar transcriptional expression of lipogenic enzymes in muscle was observed. Therefore, the type of pregnancy can explain the slight differences in mRNA expression that were found in angiogenesis and cell turnover/lactogenesis in mammary gland, although these differences not only did not affect the plasma metabolic response in ewes, but they also had no influence on plasma metabolic response, performance, and muscle transcriptional expression of their lambs.

Effect of Diet and Type of Pregnancy on Transcriptional Expression of Selected Genes in Sheep Mammary Gland.

These trials were carried out to determine firstly the effect of diet and type of pregnancy on the transcriptional expression of genes involved in angiogenesis and cell turnover/lactogenesis inside the sheep mammary gland from late gestation to late lactation. Eighteen Ile de France sheep, 8 twin- and 10 single-bearing ewes were alloted into two groups according to their diet, either based on ad libitum naturalized pasture or red clover hay plus lupine from day -45 pre-partum until day +60 post-partum. Samples from diets and mammary glands were collected at day -10 pre partum (time 1), day +30 (time 2) and day +60 post-partum (time 3) and analyzed by qRT-PCR. Additionally, samples from longissimus dorsi muscle were taken from lambs twice, at weaning and 45 days later, to determine the effect of the maternal treatment with regard to diet and type of pregnancy, on the mRNA expression of genes involved in lipid metabolism. The data was processed using the lme4 package for R, and SPSS Statistics 23.0 for Windows®. The results showed that the group of twin-bearing ewes fed red clover showed a higher expression of genes involved in angiogenesis before lambing and in cell turnover/lactogenesis during late lactation, explained by a lamb survival mechanism to delay apoptosis as a way to keep a secretory cells population and boosted by the diet quality, assuring a longer milk production potential during late lactation. Regarding lambs, apparently the maternal diet would influence the transcriptional expression of lipogenic enzymes in the longissimus dorsi muscle after weaning, but further studies are necessary to validate these results. In summary, Twin-bearing ewes fed red clover performed best at increasing the expression of genes associated with angiogenesis and cell turnover/lactogenesis in the mammary gland.

Effect of Breed on Transcriptional and Protein Expression of Lipogenic Enzymes in Tail and Subcutaneous Adipose Tissue from Two Grazing Breeds of Lambs.

This experiment was carried out to determine the effect of breed on mRNA and protein expression levels of lipogenic enzymes acetyl-CoA carboxylase α (ACC), fatty acid synthase (FAS), stearoyl-CoA desaturase 1 (SCD1) plus sterol regulatory element binding transcription factor 1c (SREBP1c) in the subcutaneous fat (SCF) from the back of the animal, and tail fat (TF) of both Chilota and Suffolk Down lambs grazing Calafatal. Eight Chilota and six Suffolk Down 2-month-old male lambs were allocated to graze a "Calafatal", a typical secondary succession of Chiloé Archipelago, Chile. After 62 d, lambs were slaughtered according to Chile's meat industry standards. Fatty acid profile, RT-qPCR, and Western blot analyses from SCF and TF samples were performed. Although the mRNA expression levels of ACC, FAS, SCD1 and SREBP1c in SCF did not differ significantly between breeds (p > 0.05), a trend to higher mRNA expression of FAS and SREBP1c in TF from Chilota lambs was observed (p = 0.06). On the other hand, FAS levels in SCF were higher in Chilota than in Suffolk Down lambs (p < 0.02), although Suffolk Down showed higher fat contents and saturated fatty acid (SFA) proportions than Chilota lambs (p < 0.01). The FAS protein expression in TF was similar in both breeds (p > 0.05). Although the fat content was higher in Suffolk Down than in Chilota lambs (p < 0.01), the SFA proportions were similar in both breeds. Finally, it can be concluded that although mRNA expression of enzymes was similar in both breeds, there were differences in some protein levels in the SCF, partially related with the fatty acid profiles, thus affecting the selection of lamb breed either for human consumption or experimental purposes.

Effects of sequential exposure to water accommodated fraction of crude oil and chlorpyrifos on molecular and biochemical biomarkers in rainbow trout.

Fish can be simultaneously or sequentially exposed to various kinds of pollutants, resulting in combined effects. Polycyclic aromatic hydrocarbons induce cytochrome P450 monooxygenase 1A (CYP1A) expression, which catalyzes the conversion of the organophosphorus insecticide chlorpyrifos (CPF) into its most active derivative, CPF-oxon. CPF-oxon inhibits CYP1A and other enzymes, including carboxylesterases (CEs) and acetylcholinesterase (AChE). We studied the effects of an in vivo exposure to crude oil water accommodated fraction (WAF) followed by an ex vivo exposure of liver tissue to CPF on the expression of Cyp1a, AhR and ARNT mRNA, CYP1A protein and on the activity of biomarker enzymes in the rainbow trout (Oncorhynchus mykiss). Juvenile rainbow trout were exposed to WAF (62 µg L-1 TPH) for 48 h. Then, liver was dissected out, sliced and exposed to 20 µg L-1 CPF ex vivo for 1 h. Liver tissue was analyzed for mRNA and protein expression and for CEs, AChE, glutathione S-transferase (GST) and CYP1A (EROD) activity. WAF induced Cyp1a mRNA and CYP1A protein expression by 10-fold and 2.5-8.3-fold, respectively, with no effect of CPF. WAF induced AhR expression significantly (4-fold) in control but not in CPF treated liver tissue. ARNT mRNA expression was significantly lowered (5-fold) by WAF. CPF significantly reduced liver EROD activity, independently of WAF pre-treatment. CEs activity was significantly inhibited in an additive manner following in vivo exposure to WAF (42%) and ex vivo exposure to CPF (19%). CPF exposure inhibited AChE activity (37%) and increased GST activity (42%).

Consecutive emamectin benzoate and deltamethrin treatments affect the expressions and activities of detoxification enzymes in the rainbow trout (Oncorhynchus mykiss).

Rainbow trout (Oncorhynchus mykiss) subjected to three consecutive, alternating treatments with emamectin benzoate (EMB) and deltamethrin (DM) during outbreaks of Caligus rogercresseyi in a farm located in southern Chile (Hornopiren, Chiloé), were studied to determine the effects of these treatments on the protein and enzymatic activity levels of cytochrome P450 1A (CYP1A), flavin-containing monooxygenase (FMO) and glutathione S-transferase (GST) in different tissues. Consecutive and alternating EMB/DM treatments resulted in a 10-fold increase and 3-fold decrease of CYP1A protein levels in the intestine and gills, respectively. Notably, CYP1A activity levels decreased in most of the analyzed tissues. FMO protein and activity levels markedly increased in the kidney and the intestine. GST was up-regulated in all tissues, either as protein or enzyme activity. When comparing consecutive EMB/DM treatments against previous studies of EMB treatment alone, CYP1A activity levels were similarly diminished, except in muscle. Likewise, FMO activity levels were increased in most of the analyzed tissues, particularly in the muscle, kidney, and intestine. The increases observed for GST were essentially unchanged between consecutive EMB/DM and EMB only treatments. These results indicate that consecutive EMB/DM treatments in rainbow trout induce the expression and activity of FMO and GST enzymes and decrease CYP1A activity. These altered activities of detoxification enzymes could generate imbalances in metabolic processes, synthesis, degradation of hormones and complications associated with drug interactions. It is especially important when analyzing possible effects of consecutive antiparasitic treatments on withholding periods and salmon farming yields.

A TRP conductance modulates repolarization after sensory-dependent depolarization in Chlamydomonas reinhardtii.

Sensory integration is vital for motile organisms constantly exposed to changing surroundings. Chlamydomonas reinhardtii is a single-celled green alga found swimming in freshwater. In this type of alga, sensory input is first detected by membrane receptors located in the cell body, and then transduced to the beating cilia by membrane depolarization. Many components of the machinery associated with sensory integration in C. reinhardtii, such as chemoreceptors and repolarization-associated channels, are yet uncharacterized. TRP channels are known mediators for cellular sensing in animal cells and it has been suggested that the C. reinhardtii genome encodes for a set of TRP proteins. Here, by combining behavioral studies with electrophysiological experiments conducted on both population and single alga, we test whether TRP channel blockers affect algal swimming behavior. Our results suggest that a TRP conductance is associated to the repolarization that follows a depolarizing receptor potential, highlighting a primitive function of TRP proteins.

A transient receptor potential ion channel in Chlamydomonas shares key features with sensory transduction-associated TRP channels in mammals.

Sensory modalities are essential for navigating through an ever-changing environment. From insects to mammals, transient receptor potential (TRP) channels are known mediators for cellular sensing. Chlamydomonas reinhardtii is a motile single-celled freshwater green alga that is guided by photosensory, mechanosensory, and chemosensory cues. In this type of alga, sensory input is first detected by membrane receptors located in the cell body and then transduced to the beating cilia by membrane depolarization. Although TRP channels seem to be absent in plants, C. reinhardtii possesses genomic sequences encoding TRP proteins. Here, we describe the cloning and characterization of a C. reinhardtii version of a TRP channel sharing key features present in mammalian TRP channels associated with sensory transduction. In silico sequence-structure analysis unveiled the modular design of TRP channels, and electrophysiological experiments conducted on Human Embryonic Kidney-293T cells expressing the Cr-TRP1 clone showed that many of the core functional features of metazoan TRP channels are present in Cr-TRP1, suggesting that basic TRP channel gating characteristics evolved early in the history of eukaryotes.