Changes in fatty acids profiles after three weeks of high-fat diet feeding in obesity-prone rats

"July 2013.""A Dissertation Presented to the Faculty of the Graduate School at the University of Missouri--Columbia In Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy."Thesis supervisor: Dr. John P. Thyfault.High-fat diet (HFD) feeding is commonly used in animal models to induce obesity and metabolic diseases. However, the effect of HFD on serum fatty acid profiles remains unclear. Changes in serum fatty acid profiles due to HFD may be a factor in lipotoxicity in various organs. Historically, in vitro studies have utilized individual non-esterified fatty acids (NEFA) to study lipid exposure, potentially ignoring the importance of fatty acid combinations on cellular lipid metabolism. OBJECTIVE: To accurately characterize the proportion of circulating fatty acids entering and exiting the liver in obese-prone rats fed with HFD in both fasted and fed state. Our ultimate goal is to create a physiological relevant fatty acid mixture to investigate lipid exposure in a cell culture system. METHODS: Obesity prone rats were fed a HFD (60%kcal fat) or a control diet (10%kcal fat) for 3 weeks; liver, portal and systemic blood samples were collected. Triglycerides (TG) and NEFA in the serum, TG, diglycerides (DG) and phospholipids (PL) in the liver were extracted and analyzed using gas chromatography. RESULTS: HFD group was heavier and had bigger fat pad compared to control diet fed animals. Both systemic and portal serum TG were ̃40% lower in HFD. In contrast, liver had higher fasting TG (̃2-fold) and DG (̃1.3-fold) in the HFD group compared to control group. Total serum NEFA levels were not affected by diet in the fasted state, but increased in the HFD group compared to the control group under fed state. In the control group, monounsaturated fatty acids (MUFA) were the predominant fatty acids in serum TG, whereas polyunsaturated fatty acids (PUFA) were the dominant fatty acids in the HFD group. The elevations of PUFA were mostly attributed to the increased of n-6 PUFA, linoleic acid and arachidonic acid. Similar shift from MUFA to n-6 PUFA also occurred in the serum NEFA fraction. CONCLUSION: HFD shifted predominate fatty acids of serum TG fraction from MUFA in the control diet to n-6 PUFA. A more physiologicIncludes bibliographical references

Effects of diet and loss of ovarian hormone production on adiposity, inflammation, cecal integrity, and gut microbial communities of rodent models

Menopause is an age-related loss of ovarian hormone production that has been linked with obesity-associated metabolic dysfunction, increased visceral adiposity, and inflammation, although the mechanisms remain unclear. Obesity has been strongly linked with profound shifts in the gastrointestinal (GI) microbiota, disrupted gut barrier function, and inflammation, but little is known regarding the involvement of the GI tract in obesity associated with loss of ovarian hormone production. Herein, we hypothesized that GI tract and the gut microbiota are involved in the weight gain and inflammation that occurs with the loss of ovarian hormone production and that dietary interventions may affect these responses. The objective of aim 1 was to evaluate the impact of soy on metabolic health, adipose tissue inflammation, and the cecal microbiota in ovariectomized (OVX) rats bred for low-running capacity (LCR), a model that has been previously shown to mimic human menopause. Forty 27-wk old LCR rats were either OVX or sham-operated (SHM) and fed either soy-rich (soy) or soy-free (control) diets for 28 wk. Soy consumption reduced (p<0.05) body weight gain, adiposity, circulating cholesterol concentrations, and improved insulin sensitivity of LCR rats. Principal coordinates analysis (PCoA) of weighted and unweighted UniFrac distances of cecal microbiota revealed a sharp separation (p<0.05) between soy- and control-fed groups. The soy-fed group had a lower (p<0.001) Firmicutes:Bacteroidetes ratio compared to control. The objective of aim 2 was to determine the energy metabolism, lipid accumulation, and inflammation in adipose and liver tissues of OVX female C57BL/6J mice in response to a high-fat diet (HFD). Forty 10-wk-old female C57BL/6J mice were fed either a high-fat diet (HFD; 60% kcal from fat) or a low-fat diet (LFD; 10% kcal from fat). After a 2-wk acclimation period, mice underwent surgical intervention (OVX or SHM). As expected, OVX mice fed HFD had substantially greater (p<0.05) body weight gain, adiposity, and hepatic triglyceride concentrations than OVX mice fed LFD. Compared to intact controls, ovariectomy led to greater (p<0.05) adipose and hepatic tissue inflammation, macrophage infiltration, oxidative stress, hindered insulin signaling and glucose uptake, and altered lipid and energy metabolism. Moreover, HFD feeding of OVX mice led to greater (p<0.05) inflammation and macrophage infiltration in gonadal adipose tissue. The degree of adiposity and inflammation resulting from HFD in OVX mice vs. SHM mice was dramatically greater than that observed in OVX mice vs. SHM control mice fed LFD. The objective of aim 3 was to examine the cecal microbial communities and barrier function in OVX or SHM mice fed a HFD or LFD for 12 wk. OVX/HFD mice had greater (p<0.05) serum lipopolysaccharide-binding protein than OVX/LFD mice. Cecal expression of inflammatory genes was not elevated due to ovariectomy, but the expression of B cell leukemia/lymphoma 2 (BCL2) was greater (p<0.05) in OVX mice than SHM mice, indicating greater apoptosis associated with loss of ovarian hormone production. Cecal permeability was not different among treatment groups. However, OVX mice had lower (p<0.05) cecal expression of occludin, claudin3, and AMP-activated protein kinase (AMPK) than SHM mice, suggesting that the cecal integrity was compromised due to loss of ovarian hormone production. Lower cecal expression of farnesoid X receptor (FXR) and fibroblast growth factor (FGF15) was observed in the OVX mice compared to SHM, suggesting an interaction between estrogen and the FXR-FGF15 pathway that is known to affect bile acid synthesis. PCoA of weighted and unweighted UniFrac distances of cecal microbiota revealed a distinct separation (p<0.05) between mice fed LFD and HFD. Despite the profound physiological changes of OVX/HFD vs. SHM/HFD mice, differential clustering of microbial communities was only observed between OVX/LFD and SHM/LFD mice. HFD promoted a greater (p<0.05) Firmicutes:Bacteroidetes ratio and lower (p<0.05) species richness of the cecal microbial community. Ovariectomy led to greater (p<0.05) abundance of Lactobacillus and lower (p<0.05) relative abundance of Oscillospira, Ruminococcus, and an undefined genus in the order Clostridiales. Differential clustering of the cecal microbial community was observed only between OVX and SHM mice fed a LFD, suggesting that the impact of ovariectomy on the cecal microbiota was masked by the HFD intervention. Thus, the objective of aim 4 was to determine the bacterial beta-glucuronidase activity level and the longitudinal shifts of the gut microbiota following the loss of ovarian function and progression of obesity in mice fed a HFD. Fecal pellets were collected at baseline (wk 0, prior to ovariectomy surgery but 2 wk after diet interventions were initiated) and 4-, 8-, and 12-wk post-surgerical intervention. Fecal beta-glucuronidase activity was elevated (p<0.05) in mice of the SHM/HFD group compared to those in the SHM/LFD group, but it was not different than mice in the OVX groups. PCoA of weighted UniFrac distances of fecal microbiota revealed a distinct separation (p<0.05) between diets. However, SHM and OVX mice only clustered differently in those fed a LFD. The Firmicutes:Bacteroidetes ratio was elevated at wk 8 and wk 12 of those fed the HFD, indicating that this elevation was due to increased adiposity instead of ovariectomy per se. The relative abundance of Clostridium and an undefined genus in the family Clostridiaceae was elevated in the OVX/HFD group at wk 4, but not SHM/HFD, indicating that this change may be due to the loss of ovarian hormone production. Our results indicate that ovariectomy impacts the composition of the gut microbial community, even though the signal was much weaker than that due to dietary intervention. However, ovariectomy promotes disrupted gut barrier function and the alteration of FXR and FGF15 in the cecum, suggesting that signaling and regulation of inflammation and bile acid metabolism in the GI tract were impacted by ovariectomy. Metagenomic, transcriptomic, and metabolomic analyses will need to be conducted to better examine the involvement of the gut microbiome and host-microbial interactions in ovariectomy- and menopausal-associated obesity

Addressing Key Risk of Water Resources and Describing the Adaptation Pathways to Uncertain Future Climate Change

Source: ICHE Conference Archive - https://mdi-de.baw.de/icheArchive

3-D neurohistology of transparent tongue in health and injury with optical clearing

Tongue receives extensive innervation to perform taste, sensory, and motor functions. Details of the tongue neuroanatomy and its plasticity in response to injury offer insights to investigate tongue neurophysiology and pathophysiology. However, due to the dispersed nature of the neural network, standard histology cannot provide a global view of the innervation. We prepared transparent mouse tongue by optical clearing to reveal the spatial features of the tongue innervation and its remodeling in injury. Immunostaining of neuronal markers, including PGP9.5 (pan-neuronal marker), calcitonin gene-related peptide (sensory nerves), tyrosine hydroxylase (sympathetic nerves), and vesicular acetylcholine transporter (cholinergic parasympathetic nerves and neuromuscular junctions), was combined with vessel painting and nuclear staining to label the tissue network and architecture. The tongue specimens were immersed in the optical-clearing solution to facilitate photon penetration for 3-dimensiontal (3-D) confocal microscopy. Taking advantage of the transparent tissue, we simultaneously revealed the tongue microstructure and innervation with subcellular-level resolution. 3-D projection of the papillary neurovascular complex and taste bud innervation was used to demonstrate the spatial features of tongue mucosa and the panoramic imaging approach. In the tongue injury induced by 4-nitroquinoline 1-oxide administration in the drinking water, we observed neural tissue remodeling in response to the changes of mucosal and muscular structures. Neural networks and the neuromuscular junctions were both found rearranged at the peri-lesional region, suggesting the nerve-lesion interactions in response to injury. Overall, this new tongue histological approach provides a useful tool for 3-D imaging of neural tissues to better characterize their roles with the mucosal and muscular components in health and disease

Role of autophagy-related proteins ATG8f and ATG8h in the maintenance of autophagic activity in Arabidopsis roots under phosphate starvation

Nutrient starvation-induced autophagy is a conserved process in eukaryotes. Plants defective in autophagy show hypersensitivity to carbon and nitrogen limitation. However, the role of autophagy in plant phosphate (Pi) starvation response is relatively less explored. Among the core autophagy-related (ATG) genes, ATG8 encodes a ubiquitin-like protein involved in autophagosome formation and selective cargo recruitment. The Arabidopsis thaliana ATG8 genes, AtATG8f and AtATG8h, are notably induced in roots under low Pi. In this study, we show that such upregulation correlates with their promoter activities and can be suppressed in the phosphate response 1 (phr1) mutant. Yeast one-hybrid analysis failed to attest the binding of the AtPHR1 transcription factor to the promoter regions of AtATG8f and AtATG8h. Dual luciferase reporter assays in Arabidopsis mesophyll protoplasts also indicated that AtPHR1 could not transactivate the expression of both genes. Loss of AtATG8f and AtATG8h leads to decreased root microsomal-enriched ATG8 but increased ATG8 lipidation. Moreover, atg8f/atg8h mutants exhibit reduced autophagic flux estimated by the vacuolar degradation of ATG8 in the Pi-limited root but maintain normal cellular Pi homeostasis with reduced number of lateral roots. While the expression patterns of AtATG8f and AtATG8h overlap in the root stele, AtATG8f is more strongly expressed in the root apex and root hair and remarkably at sites where lateral root primordia develop. We hypothesize that Pi starvation-induction of AtATG8f and AtATG8h may not directly contribute to Pi recycling but rely on a second wave of transcriptional activation triggered by PHR1 that fine-tunes cell type-specific autophagic activity