Differential Pre-mRNA Splicing Regulates Nnat Isoforms in the Hypothalamus after Gastric Bypass Surgery in Mice

Background

Neuronatin (NNAT) is an endoplasmic reticulum proteolipid implicated in intracellular signalling. Nnat is highly-expressed in the hypothalamus, where it is acutely regulated by nutrients and leptin. Nnat pre-mRNA is differentially spliced to create Nnat-α and -β isoforms. Genetic variation of NNAT is associated with severe obesity. Currently, little is known about the long-term regulation of Nnat.

Methods

Expression of Nnat isoforms were examined in the hypothalamus of mice in response to acute fast/feed, chronic caloric restriction, diet-induced obesity and modified gastric bypass surgery. Nnat expression was assessed in the central nervous system and gastrointestinal tissues. RTqPCR was used to determine isoform-specific expression of Nnat mRNA.

Results

Hypothalamic expression of both Nnat isoforms was comparably decreased by overnight and 24-h fasting. Nnat expression was unaltered in diet-induced obesity, or subsequent switch to a calorie restricted diet. Nnat isoforms showed differential expression in the hypothalamus but not brainstem after bypass surgery. Hypothalamic Nnat-β expression was significantly reduced after bypass compared with sham surgery (P = 0.003), and was positively correlated with post-operative weight-loss (R² = 0.38, P = 0.01). In contrast, Nnat-α expression was not suppressed after bypass surgery (P = 0.19), and expression did not correlate with reduction in weight after surgery (R² = 0.06, P = 0.34). Hypothalamic expression of Nnat-β correlated weakly with circulating leptin, but neither isoform correlated with fasting gut hormone levels post- surgery. Nnat expression was detected in brainstem, brown-adipose tissue, stomach and small intestine.

Conclusions

Nnat expression in hypothalamus is regulated by short-term nutrient availability, but unaltered by diet-induced obesity or calorie restriction. While Nnat isoforms in the hypothalamus are co-ordinately regulated by acute nutrient supply, after modified gastric bypass surgery Nnat isoforms show differential expression. These results raise the possibility that in the radically altered nutrient and hormonal milieu created by bypass surgery, resultant differential splicing of Nnat pre-mRNA may contribute to weight-loss.     

Seasonal variations of cellular stress response of the gilthead sea bream (Sparus aurata)

The present study aimed to investigate the seasonal cellular stress response in vital organs, like the heart, the liver, the whole blood and the skeletal (red and white) muscles of the Mediterranean fish Sparus aurata during a 1-year acclimatization period in the field, in two examined depths (0–2 m and 10–12 m). Processes studied included heat shock protein expression and protein kinase activation. Molecular responses were addressed through the expression of Hsp70 and Hsp90, the phosphorylation of stress-activated protein kinases and particularly p38 mitogen-activated protein kinase (p38 MAPK), the extracellular signal-regulated kinases (ERK-1/2) and c-Jun N-terminal kinases (JNK1/2/3). The induction of Hsp70 and Hsp90 and the phosphorylation of p38 MAPK, JNKs and ERKs in the examined five tissues of the gilthead sea bream indicated a cellular stress response under the prism of a seasonal pattern which was characterized by distinct tissue specificity. Specifically, Hsp induction and MAPK activation occurred before peak summer water temperatures, with no further increases in their levels despite increases in water temperatures. Moreover, although water temperature did not vary significantly with depth of immersion, significant effects of depth on cellular stress response were observed, probably caused by different light regime. The expression and the activation of these certain proteins can be used as tools to define the extreme thermal limits of the gilthead sea bream.     

<Emphasis Type="Italic">Thermus thermophilus</Emphasis> genome analysis: benefits and implications

The genome sequence analysis of Thermus thermophilus HB27, a microorganism with high biotechnological potential, has recently been published. In that report, the chromosomal and the megaplasmid sequence were compared to those of other organisms and discussed on the basis of their physiological and metabolic features. Out of the 2,218 putative genes identified through the large genome sequencing project, a significant number has potential interest for biotechnology. The present communication will discuss the accumulating information on molecules participating in fundamental biological processes or having potential biotechnological importance.     

Hydroxyl-platelet-activating factor exists in blood of healthy volunteers and periodontal patients

Periodontal diseases are localized chronic inflammatory conditions of the gingival and underlying bone and connective tissue. Platelet-activating factor (PAF), a potent inflammatory phospholipid mediator that has been previously detected in elevated levels in inflamed gingival tissues, in gingival crevicular fluid and in saliva, is implicated in periodontal disease. Our results from previous studies showed that the biologically active phospholipid detected in gingival crevicular fluid is a hydroxyl-PAF analogue. In this study, hydroxyl-PAF analogue was detected for the first time in human blood derived from patients with chronic periodontitis as well as from periodontally healthy volunteers. The hydroxyl-PAF analogue was purified by high-performance liquid chromatography, detected by biological assays and identified by electrospray analysis. In addition, the quantitative determination of PAF and hydroxyl-PAF analogue (expressed as PAF-like activity) showed a statistically significant increase in the ratio of hydroxyl-PAF analogue levels to PAF levels in periodontal patients, suggesting that this bioactive lipid may play a role in oral inflammation.     

Cytokine soluble receptors in perinatal and early neonatal life

BACKGROUND: In contrast to cellular receptors, soluble receptors do not enhance the cellular activation because they do not have transmembranic and cytoplasmic parts, acting thereby as endogenous regulatory mechanisms against systemic functions of cytokines. AIM: To measure serum concentrations of the soluble interleukin-2 receptor (sIL2R), soluble interleukin-4 receptor (sIL4R), soluble interleukin-6 receptor (sIL6R), and soluble tumor necrosis factor-alpha receptor I and soluble tumor necrosis factor-alpha receptor II, during the perinatal and early neonatal period, in order to evaluate their role in activation of immune response in labor and the first days postpartum. METHODS: Soluble receptor serum concentrations were determined by enzyme-linked immunosorbent assay, in 45 healthy, full-termed neonates during the first and fifth days after birth, in 25 of their mothers (MS), in 25 samples of umbilical cords (UC) and in 25 healthy adult donors age-matched with the mothers (controls). RESULTS: Soluble receptor serum concentrations showed considerable changes during labor and early neonatal life, being significantly higher both in MS (except sIL6R) and in neonatal sample UC, first and fifth days after birth, compared with controls (p<0.0001). Neonatal serum sIL2R and sIL6R increased significantly from birth to the fifth day, while the remaining receptors showed a rapid increase in the first day (p<0.0001), declining significantly thereafter (p<0.0001). CONCLUSION: Our findings suggest that the elevated concentrations of all studied soluble cytokine receptors reflect the activation of immune response, and represent also regulatory protective mechanisms for mother and fetus-neonate against the systemic function of cytokines during labor and early neonatal life.     

β-Adrenergic Receptors Mediate a Stress-Induced Decrease in IGF-II mRNA in the Rat Cerebellum

1. Exposure to a combined forced swimming–confinement stress resulted in a decrease in insulin-like growth factor II (IGF-II) mRNA levels in the whole brain (without the cerebellum) and in the isolated brain areas of the cerebral cortex, the hippocampus, and the cerebellum.2. In an effort to elucidate the neurotransmitter systems involved in this stress-induced decrease, animals were injected prior to exposure to the stress, with either propranolol, diazepam, or MK-801.3. Administration of diazepam or MK-801 did not affect the stress-induced decrease in IGF-II mRNA in any of the three brain areas examined.4. Administration of propranolol prior to the exposure to the stress inhibited the stress-induced decrease in IGF-II mRNA in the cerebellum. Propranolol had no such effect in the cerebral cortex or the hippocampus.5. Our results suggest that in the cerebellum, the stress-induced decrease in IGF-II mRNA is mediated by ₂-adrenergic receptors.     

Adult Consequences of Post-weaning High Fat Feeding on the Limbic–HPA Axis of Female Rats

The peripubertal period is critical for the final maturation of circuits controlling energy homeostasis and stress response. However, the consequence of juvenile fat consumption on adult physiology is not clear. This study analyzed the adult consequences of post-weaning fat feeding on limbic–hypothalamic–pituitary–adrenal (HPA) axis components and on metabolic regulators of female rats. Wistar rats were fed either a high fat (HF) diet or the normal chow from weaning to puberty or to 3 months of age. Additional groups crossed their diets at puberty onset. Plasma leptin, insulin, and corticosterone levels were determined by radioimmunoassay and their brain receptors by western blot analysis. Adult HF-fed animals though not overweight, had higher corticosterone and reduced glucocorticoid receptor levels in the hypothalamus and hippocampus, compared to the controls. The alterations in HPA axis emerged already at puberty onset. Leptin receptor levels in the hypothalamus were reduced only by continuous fat feeding from weaning to adulthood. The pre-pubertal period appeared more vulnerable to diet-induced alterations in adulthood than the post-pubertal one. Switching from fat diet to normal chow at puberty onset restored most of the diet-induced alterations in the HPA axis. The corticosteroid circuit rather than the leptin or insulin system appears as the principal target for the peripubertal fat diet-induced effects in adult female rats.     

Photosystem 2 activity of <Emphasis Type="Italic">Citrus volkameriana</Emphasis> (L.) leaves as affected by Mn nutrition and irradiance

Citrus volkameriana (L.) plants were grown for 43 d in nutrient solutions containing 0, 2, 14, 98, or 686 μM Mn (Mn₀, Mn₂, Mn₁₄, Mn₉₈, and Mn₆₈₆, respectively). To adequately investigate the combined effects of Mn nutrition and irradiance on photosystem 2 (PS2) activity, irradiance response curves for electron transport rate (ETR), nonphotochemical quenching (q_N), photochemical quenching (q_P), and real photochemical efficiency of PS2 (Φ_PS2) were recorded under 10 different irradiances (66, 96, 136, 226, 336, 536, 811, 1 211, 1 911, and 3 111 μmol m⁻² s⁻¹, I₆₆ to I₃₁₁₁, respectively) generated with the PAM-2000 fluorometer. Leaf chlorophyll content was significantly lower under Mn excess (Mn₆₈₆) compared to Mn₀; its highest values were recorded in the treatments Mn₂-Mn₉₈. However, ETR and Φ_PS2 values were significantly lower under Mn₀ compared to the other Mn treatments, when plants were exposed to irradiances ≥96 μmol m⁻² s⁻¹. Furthermore, Mn₀ plants had significantly higher values of q_N and lower values of q_P at irradiances ≤226 and ≥336 μmol m⁻² s⁻¹, respectively, than those grown under Mn₂-Mn₆₈₆. Irrespective of Mn treatment, the values of Φ_PS2 and q_N decreased, while those of q_P increased progressively by increasing irradiance from I₁₃₆ to I₃₁₁₁. Finally, Mn₂-Mn₉₈ plants were less sensitive to photoinhibition of photosynthesis (≥811 μmol m⁻² s⁻¹) than the Mn₆₈₆ (≥536 μmol m⁻² s⁻¹) and Mn₀ (≥336 μmol m⁻² s⁻¹) ones.