High Oestrogen receptor alpha expression correlates with adverse prognosis and promotes metastasis in colorectal cancer.

In normal colon tissue, oestrogen receptor alpha (ERα) is expressed at low levels, while oestrogen receptor beta (ERß) is considered the dominant subtype. However, in colon carcinomas, the ERα/ß ratio is often increased, an observation that prompted us to further investigate ERα's role in colorectal cancer (CRC). Here, we assessed ERα nuclear expression in 351 CRC patients. Among them, 119 exhibited positive ERα nuclear expression, which was significantly higher in cancer tissues than in matched normal tissues. Importantly, patients with positive nuclear ERα expression had a poor prognosis. Furthermore, positive ERα expression correlated with increased levels of the G-protein coupled cysteinyl leukotriene receptor 1 (CysLT1R) and nuclear ß-catenin, both known tumour promoters. In mouse models, ERα expression was decreased in Cysltr1-/- CAC (colitis-associated colon cancer) mice but increased in ApcMin/+ mice with wild-type Cysltr1. In cell experiments, an ERα-specific agonist (PPT) increased cell survival via WNT/ß-catenin signalling. ERα activation also promoted metastasis in a zebrafish xenograft model by affecting the tight junction proteins ZO-1 and Occludin. Pharmacological blockade or siRNA silencing of ERα limited cell survival and metastasis while restoring tight junction protein expression. In conclusion, these findings highlight the potential of ERα as a prognostic marker for CRC and its role in metastasis.

GFAP hyperpalmitoylation exacerbates astrogliosis and neurodegenerative pathology in PPT1-deficient mice.

The homeostasis of protein palmitoylation and depalmitoylation is essential for proper physiological functions in various tissues, in particular the central nervous system (CNS). The dysfunction of PPT1 (PPT1-KI, infantile neuronal ceroid lipofuscinosis [INCL] mouse model), which catalyze the depalmitoylation process, results in serious neurodegeneration accompanied by severe astrogliosis in the brain. Endeavoring to determine critical factors that might account for the pathogenesis in CNS by palm-proteomics, glial fibrillary acidic protein (GFAP) was spotted, indicating that GFAP is probably palmitoylated. Questions concerning if GFAP is indeed palmitoylated in vivo and how palmitoylation of GFAP might participate in neural pathology remain unexplored and are waiting to be investigated. Here we show that GFAP is readily palmitoylated in vitro and in vivo; specifically, cysteine-291 is the unique palmitoylated residue in GFAP. Interestingly, it was found that palmitoylated GFAP promotes astrocyte proliferation in vitro. Furthermore, we showed that PPT1 depalmitoylates GFAP, and the level of palmitoylated GFAP is overwhelmingly up-regulated in PPT1-knockin mice, which lead us to speculate that the elevated level of palmitoylated GFAP might accelerate astrocyte proliferation in vivo and ultimately led to astrogliosis in INCL. Indeed, blocking palmitoylation by mutating cysteine-291 into alanine in GFAP attenuate astrogliosis, and remarkably, the concurrent neurodegenerative pathology in PPT1-knockin mice. Together, these findings demonstrate that hyperpalmitoylated GFAP plays critical roles in regulating the pathogenesis of astrogliosis and neurodegeneration in the CNS, and most importantly, pinpointing that cysteine-291 in GFAP might be a valuable pharmaceutical target for treating INCL and other potential neurodegenerative diseases.

Manual Therapy Improves Fibromyalgia Symptoms by Downregulating SIK1.

Fibromyalgia (FM), classified by ICD-11 with code MG30.0, is a chronic debilitating disease characterized by widespread pain, fatigue, cognitive impairment, sleep, and intestinal alterations, among others. FM affects a large proportion of the worldwide population, with increased prevalence among women. The lack of understanding of its etiology and pathophysiology hampers the development of effective treatments. Our group had developed a manual therapy (MT) pressure-controlled custom manual protocol on FM showing hyperalgesia/allodynia, fatigue, and patient's quality of life benefits in a cohort of 38 FM cases (NCT04174300). With the aim of understanding the therapeutic molecular mechanisms triggered by MT, this study interrogated Peripheral Blood Mononuclear Cell (PBMC) transcriptomes from FM participants in this clinical trial using whole RNA sequencing (RNAseq) and reverse transcription followed by quantitative Polymerase Chain Reaction (RT-qPCR) technologies. The results show that the salt-induced kinase SIK1 gene was consistently downregulated by MT in FM, correlating with improvement of patient symptoms. In addition, this study compared the findings in a non-FM control cohort subjected to the same MT protocol, evidencing that those changes in SIK1 expression with MT only occurred in individuals with FM. This positions SIK1 as a potential biomarker to monitor response to MT and as a therapeutic target of FM, which will be further explored by continuation studies.

High PPT1 expression predicts poor clinical outcome and PPT1 inhibitor DC661 enhances sorafenib sensitivity in hepatocellular carcinoma.

BACKGROUND: Adaptive resistance and side effects of sorafenib treatment result in unsatisfied survival of patients with hepatocellular carcinoma (HCC). Palmitoyl-protein thioesterase 1 (PPT1) plays a critical role in progression of various cancers. However, its role on prognosis and immune infiltrates in HCC remains unclarified. METHODS: By data mining in the Cancer Genome Atlas databases, the role of PPT1 in HCC were initially investigated. Furthermore, HCC cell lines Hep 3B and Hep 1-6 were treated with DC661 or siRNA against PPT1. The biological function of PPT1 was determined by CCK-8 test, colony formation assay, TUNEL staining, immunofluorescence staining, Western blot test, and PI-Annexin V apoptosis assays in vitro. Animal models of subcutaneous injection were applied to investigate the therapeutic role of targeting PPT1. RESULTS: We found that PPT1 levels were significantly upregulated in HCC tissues compared with normal tissues and were significantly associated with a poor prognosis. Multivariate analysis further confirmed that high expression of PPT1 was an independent risk factor for poor overall survival of HCC patients. We initially found that PPT1 was significantly upregulated in sorafenib-resistant cell lines established in this study. Upon sorafenib treatment, HCC cells acquired adaptive resistance by inducing autophagy. We found that DC661, a selective and potent small-molecule PPT1-inhibitor, induced lysosomal membrane permeability, caused lysosomal deacidification, inhibited autophagy and enhanced sorafenib sensitivity in HCC cells. Interestingly, this sensitization effect was also mediated by the induction mitochondrial pathway apoptosis. In addition, the expression level of PPT1 was associated with the immune infiltration in the HCC tumor microenvironment, and PPT1 inhibitor DC661 significantly enhanced the anti-tumor immune response by promoting dendritic cell maturation and further promoting CD8+ T cell activation. Moreover, DC661 combined with sorafenib was also very effective at treating tumor models in immunized mice. CONCLUSIONS: Our findings suggest that targeting PPT1 with DC661 in combination with sorafenib might be a novel and effective alternative therapeutic strategy for HCC.

Seizures in PPT1 Knock-In Mice Are Associated with Inflammatory Activation of Microglia.

Infantile neuronal ceroid lipofuscinosis (INCL), the most severe form of neuronal ceroid lipofuscinoses, is caused by mutations in the lysosomal enzyme palmitoyl protein thioesterase 1 (PPT1). Typical symptoms of this disease include progressive psychomotor developmental retardation, visual failure, seizures, and premature death. Here, we investigated seizure activity and relevant pathological changes in PPT1 knock-in mice (PPT1 KI). The behavior studies in this study demonstrated that PPT1 KI mice had no significant seizure activity until 7 months of age, and local field potentials also displayed epileptiform activity at the same age. The expression levels of Iba-1 and CD68 demonstrated, by Western blot analysis, the inflammatory cytokine TNF-α content measured with enzyme-linked immunosorbent assay, and the number of microglia demonstrated by immunohistochemistry (IHC) were significantly increased at age of 7 months, all of which indicate microglia activation at an age of seizure onset. The increased expression of GFAP were seen at an earlier age of 4 months, and such an increase reached its peak at age of 6 months, indicating that astrocyte activation precedes microglia. The purinergic P2X7 receptor (P2X7R) is an ATP-sensitive ionic channel that is highly expressed in microglia and is fundamental to microglial activation, proliferation, cytokines release and epilepsy. We show that the ATP concentration in hippocampal tissue in PPT1 KI mice was increased using an enhanced ATP assay kit and demonstrated that the antagonist of P2X7R, A-438079, significantly reduced seizures in PPT1 KI mice. In contrast to glial cell activation and proliferation, a significant reduction in synaptic proteins GABAAR was seen in PPT1 KI mice. These results indicate that seizure in PPT1 KI mice may be associated with microglial activation involved in ATP-sensitive P2X7R signaling and impaired inhibitory neurotransmission.

A case of juvenile CLN1- challenge in diagnosis and epilepsy treatment.

IntroductionClassic onset of CLN1 disease is within the first year of life with developmental arrest, epilepsy and rapid progression. In an atypical variant of CLN1 disease onset is later in the juvenile epoch. Although epilepsy in the juvenile form of CLN1 often is less severe than in typical CLN1, treatment of seizures and status epilepticus may be challenging.Case presentationThe clinical course, misdiagnosis and epilepsy phenotype are presented in a girl with juvenile CLN1. Cognitive and neurologic regression started at age 5.5 years. Epilepsy was a major clinical issue as the patient suffered from focal seizures, recurrent status epilepticus and epilepsia partialis continua. In one episode of refractory status epilepticus, the patient had significant bradycardia associated with the intravenous infusion of levetiracetam. Diagnosis was made at the age of 12 years, based on palmitoyl protein-thioesterase (PPT) enzyme deficiency and genetic testing that documented a homozygous exon missense mutation in the CLN1 gene (PPT1, c.541G>A, p.Val181Met).DiscussionEpilepsy in all NCL patients is a major clinical issue and presumed related to neuronal excitation and epileptogenesis. The treatment of status epilepticus, in juvenile CLN1 patients, presents a particular challenge and requires monitoring of potential serious pharmacologic side effects of therapy.

Current state of diagnostic, screening and surveillance testing methods for COVID-19 from an analytical chemistry point of view.

Since December 2019, we have been in the battlefield with a new threat to the humanity known as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In this review, we describe the four main methods used for diagnosis, screening and/or surveillance of SARS-CoV-2: Real-time reverse transcription polymerase chain reaction (RT-PCR); chest computed tomography (CT); and different complementary alternatives developed in order to obtain rapid results, antigen and antibody detection. All of them compare the highlighting advantages and disadvantages from an analytical point of view. The gold standard method in terms of sensitivity and specificity is the RT-PCR. The different modifications propose to make it more rapid and applicable at point of care (POC) are also presented and discussed. CT images are limited to central hospitals. However, being combined with RT-PCR is the most robust and accurate way to confirm COVID-19 infection. Antibody tests, although unable to provide reliable results on the status of the infection, are suitable for carrying out maximum screening of the population in order to know the immune capacity. More recently, antigen tests, less sensitive than RT-PCR, have been authorized to determine in a quicker way whether the patient is infected at the time of analysis and without the need of specific instruments.

Investigating the Endo-Lysosomal System in Major Neurocognitive Disorders Due to Alzheimer's Disease, Frontotemporal Lobar Degeneration and Lewy Body Disease: Evidence for SORL1 as a Cross-Disease Gene.

Dysfunctions in the endo-lysosomal system have been hypothesized to underlie neurodegeneration in major neurocognitive disorders due to Alzheimer's disease (AD), Frontotemporal Lobar Degeneration (FTLD), and Lewy body disease (DLB). The aim of this study is to investigate whether these diseases share genetic variability in the endo-lysosomal pathway. In AD, DLB, and FTLD patients and in controls (948 subjects), we performed a targeted sequencing of the top 50 genes belonging to the endo-lysosomal pathway. Genetic analyses revealed (i) four previously reported disease-associated variants in the SORL1 (p.N1246K, p.N371T, p.D2065V) and DNAJC6 genes (p.M133L) in AD, FTLD, and DLB, extending the previous knowledge attesting SORL1 and DNAJC6 as AD- and PD-related genes, respectively; (ii) three predicted null variants in AD patients in the SORL1 (p.R985X in early onset familial AD, p.R1207X) and PPT1 (p.R48X in early onset familial AD) genes, where loss of function is a known disease mechanism. A single variant and gene burden analysis revealed some nominally significant results of potential interest for SORL1 and DNAJC6 genes. Our data highlight that genes controlling key endo-lysosomal processes (i.e., protein sorting/transport, clathrin-coated vesicle uncoating, lysosomal enzymatic activity regulation) might be involved in AD, FTLD and DLB pathogenesis, thus suggesting an etiological link behind these diseases.

EVDHM-ARIMA-Based Time Series Forecasting Model and Its Application for COVID-19 Cases.

The time-series forecasting makes a substantial contribution in timely decision-making. In this article, a recently developed eigenvalue decomposition of Hankel matrix (EVDHM) along with the autoregressive integrated moving average (ARIMA) is applied to develop a forecasting model for nonstationary time series. The Phillips-Perron test (PPT) is used to define the nonstationarity of time series. EVDHM is applied over a time series to decompose it into respective subcomponents and reduce the nonstationarity. ARIMA-based model is designed to forecast the future values for each subcomponent. The forecast values of each subcomponent are added to get the final output values. The optimized value of ARIMA parameters for each subcomponent is obtained using a genetic algorithm (GA) for minimum values of Akaike information criterion (AIC). Model performance is evaluated by estimating the future values of daily new cases of the recent pandemic disease COVID-19 for India, USA, and Brazil. The high efficacy of the proposed method is convinced with the results.

Strategies for Positive Partial Transpose (PPT) States in Quantum Metrologies with Noise.

Quantum metrology overcomes standard precision limits and has the potential to play a key role in quantum sensing. Quantum mechanics, through the Heisenberg uncertainty principle, imposes limits on the precision of measurements. Conventional bounds to the measurement precision such as the shot noise limit are not as fundamental as the Heisenberg limits, and can be beaten with quantum strategies that employ 'quantum tricks' such as squeezing and entanglement. Bipartite entangled quantum states with a positive partial transpose (PPT), i.e., PPT entangled states, are usually considered to be too weakly entangled for applications. Since no pure entanglement can be distilled from them, they are also called bound entangled states. We provide strategies, using which multipartite quantum states that have a positive partial transpose with respect to all bi-partitions of the particles can still outperform separable states in linear interferometers.

Comparison of home-based pencil push-up therapy and office-based orthoptic therapy in symptomatic patients of convergence insufficiency: a randomized controlled trial.

PURPOSE: To assess and compare the effectiveness of home-based pencil push-up therapy (PPT) and office-based orthoptic therapy (OBOT) in patients with convergence insufficiency. METHODS: In this randomized clinical trial, 176 symptomatic patients with convergence insufficiency, aged between 9 to 30 years, were randomly assigned to receive 6 weeks of home-based PPT (Group I) or OBOT (Group II) after determining refractive error, near point of convergence (NPC), convergence insufficiency symptom survey (CISS) score, near phoria and positive fusional vergences (PFV) at near. The participants of Group I underwent home-based PPT (pencil push-ups exercises15 minutes per day, daily for 6 weeks) and those of Group II OBOT (convergence fusional exercises on synoptophore for 20 min per day, 3 days a week, for 6 weeks) without home reinforcement. Patients were re-examined at 3 and 6 weeks after initiation of treatment. NPC and CISS score were the primary and secondary outcome measures, respectively. Statistical analysis was performed with the independent samples t-test, Friedman test and the analysis of variance (ANOVA). Statistical significance was indicated by p-value < 0.05. RESULTS: Participants of both the groups had statistically significant improvement in NPC, CISS score, PFV and near phoria (p < 0.001), but there was no statistically significant difference between the two groups (p > 0.05). However, patients of Group II had significantly better PFV after final visit than those of Group I (p < 0.001). CONCLUSION: Home-based PPT with good suppression control and with compliance ensured by log book entries, is a simple, cheap, less time consuming and comparably effective alternative to more expensive OBOT for patients suffering from CI. CTRI registration number: REF/2016/11/012,732, Date of registration 25/04/ 2016, Retrospectively Registered.

Pressure pain thresholds in office workers with chronic neck pain: A systematic review and meta-analysis.

OBJECTIVES: The purpose of this study was to (a) compare pressure pain threshold (PPT) values between office workers with chronic neck pain and asymptomatic controls; (b) establish reference PPT values in chronic neck pain; and (c) evaluate associations between PPTs and pain intensity, and disability. METHODS: Seven English/Portuguese databases were searched for relevant literature. Studies investigating adult office workers (age >18 years) with chronic neck pain were included if PPTs were an outcome. The risk of bias was assessed using the Downs and Black checklist. Meta-analysis was conducted if a cluster contained at least two studies reporting the same PPTs. RESULTS: Ten high quality, two low quality, and one poor quality studies were included. The meta-analysis revealed decreased PPT values in the upper trapezius, extensor carpi ulnaris, and tibialis anterior in office workers with chronic neck pain when compared with healthy workers, without a statistical difference (p > 0.05). The PPT reference value in the upper trapezius was 263 kPa (95% confidence interval [CI] = 236.35 to 289.70), and 365 kPa (95% CI = 316.66 to 415.12) for the tibialis anterior in office workers with chronic neck pain. No correlations were found between the upper trapezius PPT and pain intensity and disability. CONCLUSION: This meta-analysis found that all the PPT measurements were not significantly reduced in office workers with chronic neck pain compared with healthy workers. These assumptions were based on a small sample of existing studies, and therefore further studies are necessary to quantify the differences in PPTs. Hypersensitivity PPT reference values are proposed for localized and extrasegmental sites in office workers with chronic neck pain.

Key role of estrogen receptor ß in the organization of brain and behavior of the Japanese quail.

Estrogens play a key role in the sexual differentiation of the brain and behavior. While early estrogen actions exert masculinizing effects on the brain of male rodents, a diametrically opposite effect is observed in birds where estrogens demasculinize the brain of females. Yet, the two vertebrate classes express similar sex differences in the brain and behavior. Although ERα is thought to play a major role in these processes in rodents, the role of ERß is still controversial. In birds, the identity of the estrogen receptor(s) underlying the demasculinization of the female brain remains unclear. The aim of the present study was thus to determine in Japanese quail the effects of specific agonists of ERα (propylpyrazole triol, PPT) and ERß (diarylpropionitrile, DPN) administered at the beginning of the sensitive period (embryonic day 7, E7) on the sexual differentiation of male sexual behavior and on the density of vasotocin-immunoreactive (VT-ir) fibers, a known marker of the organizational action of estrogens on the quail brain. We demonstrate that estradiol benzoate and the ERß agonist (DPN) demasculinize male sexual behavior and decrease the density of VT-ir fibers in the medial preoptic nucleus and the bed nucleus of the stria terminalis, while PPT has no effect on these measures. These results clearly indicate that ERß, but not ERα, is involved in the estrogen-induced sexual differentiation of brain and sexual behavior in quail.

Estrogen receptor α activation aggravates imiquimod-induced psoriasis-like dermatitis in mice by enhancing dendritic cell interleukin-23 secretion.

Our recent study has reported that estrogen receptors (ERs) are involved in several types of allergy development. This study aims to investigate the possible relationship between ER activation and development of imiquimod-induced psoriasis-like dermatitis. A mouse model of imiquimod-induced psoriasis-like dermatitis was generated by 5 days of topical application of 5% of imiquimod cream on the back of the ear and the shaved back skin of male BALB/c mice. From the second day of applying 5% imiquimod cream, either ERα selective agonist (propylpyrazoletriol [PPT] 2.5 mg/kg) or ERß selective agonist (diarylpropionitrile, DPN; 2.5 mg/kg) was administered orally for four consecutive days. Immediately after the final imiquimod cream application, scratching behavior was video monitored for 2 hours. The ear-swelling response was determined by comparing ear thickness before and after the final application of imiquimod cream. Twenty-four hours after the final imiquimod application, back skin tissue and auricular lymph nodes were isolated under isoflurane anesthesia. Oral administration of PPT significantly induced itch behavior and proinflammatory responses, including the levels of interleukin (IL)-17 and IL-22, whereas DPN treatment did not influence either pruritic or proinflammatory responses. In addition, IL-23 contribution by dendritic cells was identified using ER agonists on pretreated lipopolysaccharide (LPS)-stimulated murine bone marrow derived dendritic cells (BMDCs). PPT also significantly enhanced IL-23 secretion by LPS-stimulated BMDCs. Our findings indicate that the activation of ERα, but not ERß, is directly associated with inflammatory and pruritic responses in a mouse model of the imiquimod-induced psoriasis by enhancing the secretion of IL-23 by dendritic cells.

Protein precipitation method for determination of clobazam and N-desmethylclobazam in human plasma by LC-MS/MS.

A protein precipitation method for the determination of clobazam (CLB) and its major active metabolite N-desmethylclobazam (N-CLB) in human plasma by liquid chromatography tandem mass spectrometry (LC-MS/MS) was established. CLB and N-CLB were extracted from human plasma samples by protein precipitation with methanol. Analyte separation was done using a Phenomenex Kinetex™ Biphenyl (50 × 2.1 mm, 1.7 µm) column using isocratic elution with a mobile phase of 5 mm ammonium formate with 0.01% ammonium hydroxide (40%) and methanol (60%) at a flow rate of 0.4 mL/min and an injection volume of 10 µL. The detection was performed on a triple quadrupole mass spectrometer in multiple reaction monitoring mode to monitor the precursor-to-product ion transitions of m/z 301.1 → 259.0, 306.0 → 263.9 for CLB and CLB-D5 and 287.0 → 245.0, 292.0 → 250.0 for N-CLB and N-CLB-D5 in positive electrospray ionization mode, respectively. The method was validated over a concentration range of 2.0-750 ng/mL for CLB and 0.7-200 ng/mL for N-CLB on SCIEX Triple Quad 4500 MS System. Total run time was 5 min. This method has been designed for bioequivalence study for formulations containing 20 mg of CLB.

Functional importance of palmitoyl protein thioesterase 1 (PPT1) expression by Sertoli cells in mediating cholesterol metabolism and maintenance of sperm quality.

Sertoli cells are a type of nurse cell in the seminiferous epithelium that are crucial for sustaining spermatogenesis by extending nutritional and energy support to the developing germ cells. Dysfunction of Sertoli cells could cause disordered spermatogenesis and reduced fertility in males. In this study, we focused on the expression and function of palmitoyl protein thioesterase 1 (PPT1), a lysosomal depalmitoylating enzyme, in Sertoli cells. Here, we show that PPT1 expression in Sertoli cells is responsive to cholesterol treatment and that specific knockout of Ppt1 in Sertoli cells causes male subfertility associated with poor sperm quality and a high ratio of sperm deformity. Specifically, Ppt1 deficiency leads to poor cell variably accompanied with abnormal lysosome accumulation and increased cholesterol levels in Sertoli cells. Further, Ppt1 deficiency results in poor adhesion of developing germ cells to Sertoli cells in the seminiferous epithelium, which is likely to be responsible for the reduced male fertility as a consequence of declines in sperm count and motility as well as a high incidence of sperm head deformity. In summary, PPT1 affects sperm quality and male fertility through regulating lysosomal function and cholesterol metabolism in Sertoli cells.

Effect of pre-exercise ingestion of α-lactalbumin on subsequent endurance exercise performance and mood states.

This study investigated the effect of pre-exercise α-lactalbumin ingestion on subsequent endurance exercise performance, muscle pain and mood states. In a two-stage cross-over counterbalance design, eleven male endurance runners (age: 31 (se 2) years, height: 169·5 (se 4·4) cm, weight: 63·6 (se 5·1) kg, V̇O2max: 58·8 (se 6·3) ml/kg per min) consumed two solutions (carbohydrate+α-lactalbumin, CA; carbohydrate+whey protein isolate, CW) 2 h before a self-paced 21-km run. Creatine kinase, IL-6, muscle pain, pressure pain threshold (PPT) and mood states were assessed 2 h before exercise, immediately before exercise (Pre-ex0) and immediately after exercise (Post-ex0). No difference was found in 21-km running performance between two trials (CA v. CW: 115·85 (se 5·20) v. 118·85 (se 5·51) min, P=0·48). Compared with CW, CA led to higher PPT at Pre-ex0 (41·77 (se 2·27) v. 35·56 (se 2·10) N/cm2, P<0·01) and Post-ex0 (38·76 (se 3·23) v. 35·30 (se 3·55) N/cm2, P=0·047). Compared with CW, CA reduced the feeling of fatigue at Post-ex0 (P<0·01); CA also reduced salivary cortisol levels at Post-ex0 (0·72 (se 0·07) v. 0·83 (se 0·13) ng/ml, P<0·01). In conclusion, the ingestion of α-lactalbumin did not improve the 21-km time-trial performance. However, compared with the pre-exercise ingestion of whey protein, that of α-lactalbumin led to superior results during similar levels of endurance exercise: it elevated PPT and reduced the feeling of fatigue and the cortisol levels.

Comparison of the growth performance and long-chain PUFA biosynthetic ability of the genetically improved farmed tilapia (Oreochromis niloticus) reared in different salinities.

To compare the growth and biosynthetic ability of long-chain PUFA (LC-PUFA) of the genetically improved farmed tilapia (GIFT) (Oreochromis niloticus) in different water salinities, an 8-week feeding trial was conducted on the GIFT juveniles at 0, 12 and 24 ‰ (parts per thousand; ppt), respectively, with three isonitrogenous (32 %) and isolipidic (8 %) diets (D1-D3). Diet D1 with fish oils (rich in LC-PUFA) as lipid source was used as the control, while D2 and D3 with vegetable oil (free LC-PUFA) blends as lipid source contained different ratios of linoleic acid (LA, 18 : 2n-6) and α-linolenic acid (ALA, 18 : 3n-3) at 4·04 (D2) and 0·54 (D3), respectively. At the end of feeding trial, the growth performance of D2 and D3 groups under all salinity treatments was as good as that of D1 group, which indicates that the GIFT juveniles may convert dietary LA and ALA into LC-PUFA to meet the requirement of essential fatty acids for normal growth and physiology. When fed the same diets, GIFT at 12 ppt had a better growth performance coupled with a higher liver and muscle arachidonic acid content than those in freshwater. Furthermore, brackish water (24 ppt) significantly promoted the mRNA levels of elongase 5 of very long-chain fatty acids (elovl5) and peroxisome proliferator-activated receptor α (pparα) in liver, when compared with freshwater. These results suggest that the GIFT may display better growth performance together with a relatively higher endogenous LC-PUFA biosynthetic ability under brackish water (12 and 24 ppt), probably through improving the expression of elovl5 and pparα in liver.

The protein phosphatase gene MaPpt1 acts as a programmer of microcycle conidiation and a negative regulator of UV-B tolerance in Metarhizium acridum.

The Ser/Thr protein phosphatase Ppt1 (yeast)/PP5 (humans) has been implicated in signal transduction-mediated growth and differentiation, DNA damage/repair, cell cycle progression, and heat shock responses. Little, however, is known concerning the functions of Ppt1/PP5 in filamentous fungi. In this study, the Ppt1 gene MaPpt1 was characterized in the insect pathogenic fungus, Metarhizium acridum. The MaPpt1 protein features a three-tandem tetratricopeptide repeat (TPR) domain and a peptidyl-prolyl cis-trans isomerase-like (PP2Ac) domain. Subcellular localization using an MaPpt1::eGFP fusion protein revealed that MaPpt1 was localized in the cytoplasm of spores, but gathered at the septa in growing hyphae. Targeted gene inactivation of MaPpt1 in M. acridum resulted in unexpected reprogramming of normal aerial conidiation to microcycle conidiation. Although overall vegetative growth was unaffected, a significant increase in conidial yield was noted in ΔMaPpt1. Stress-responsive phenotypes and virulence were largely unaffected in ΔMaPpt1. Exceptionally, ΔMaPpt1 displayed increased UV tolerance compared to wild type. Digital gene expression data revealed that MaPpt1 mediates transcription of sets of genes involved in conidiation, polarized growth, cell cycle, cell proliferation, DNA replication and repair, and some important signaling pathways. These data indicate a unique role for Ppt1 in filamentous fungal development and differentiation.

Cholinergic, Glutamatergic, and GABAergic Neurons of the Pedunculopontine Tegmental Nucleus Have Distinct Effects on Sleep/Wake Behavior in Mice.

The pedunculopontine tegmental (PPT) nucleus has long been implicated in the regulation of cortical activity and behavioral states, including rapid eye-movement (REM) sleep. For example, electrical stimulation of the PPT region during sleep leads to rapid awakening, whereas lesions of the PPT in cats reduce REM sleep. Though these effects have been linked with the activity of cholinergic PPT neurons, the PPT also includes intermingled glutamatergic and GABAergic cell populations, and the precise roles of cholinergic, glutamatergic, and GABAergic PPT cell groups in regulating cortical activity and behavioral state remain unknown. Using a chemogenetic approach in three Cre-driver mouse lines, we found that selective activation of glutamatergic PPT neurons induced prolonged cortical activation and behavioral wakefulness, whereas inhibition reduced wakefulness and increased non-REM (NREM) sleep. Activation of cholinergic PPT neurons suppressed lower-frequency electroencephalogram rhythms during NREM sleep. Last, activation of GABAergic PPT neurons slightly reduced REM sleep. These findings reveal that glutamatergic, cholinergic, and GABAergic PPT neurons differentially influence cortical activity and sleep/wake states. SIGNIFICANCE STATEMENT: More than 40 million Americans suffer from chronic sleep disruption, and the development of effective treatments requires a more detailed understanding of the neuronal mechanisms controlling sleep and arousal. The pedunculopontine tegmental (PPT) nucleus has long been considered a key site for regulating wakefulness and REM sleep. This is mainly because of the cholinergic neurons contained in the PPT nucleus. However, the PPT nucleus also contains glutamatergic and GABAergic neurons that likely contribute to the regulation of cortical activity and sleep-wake states. The chemogenetic experiments in the present study reveal that cholinergic, glutamatergic, and GABAergic PPT neurons each have distinct effects on sleep/wake behavior, improving our understanding of how the PPT nucleus regulates cortical activity and behavioral states.