The inverted U-shaped relationship between epinephrine and pancreatic ductal adenocarcinoma patients' survival with compensation of lymphocyte.

BACKGROUND: The relationship between epinephrine and cancer can be dose-dependent in in vivo study. Whether it is the same in human body still needs verification. METHOD: We used frozen human pancreatic ductal adenocarcinoma (PDAC) tissues to detect epinephrine content and analyzed its relationship with survival using the K-M method and Cox regression. Disturbance of blood cell count and C-reactive protein and identification of related potent intermediary factors were also analyzed. RESULTS: K-M plot and Cox regression all showed the inverted U-shaped relationship between epinephrine and PDAC survival. Lymphocyte adjustment can increase the HRs of epinephrine for PDAC death by >10%. CONCLUSION: Epinephrine played an anti-tumor or pro-tumor effect depending on the specific concentration. Circulating lymphocyte count was elevated and might acted as a compensation pathway to reduce the pro-tumor effect of epinephrine to PDAC.

Revision of the subgenus Stegana (Steganina) from China, with assessment of species delimitation using DNA barcodes (Diptera, Drosophilidae).

A total of 58 (eight known and 50 new) species of the subgenus Stegana (Steganina) from China were surveyed and (re)described: S. (S.) bacilla Chen & Aotsuka, 2004, S. (S.) belokobylskiji Sidorenko, 1997, S. (S.) hirticeps Wang, Gao, & Chen, 2013, S. (S.) izu Sidorenko, 1997, S. (S.) kanmiyai Okada & Sidorenko, 1992, S. (S.) masanoritodai Okada & Sidorenko, 1992, S. (S.) maymyo Sidorenko, 1997, stat. rev., S. (S.) nigripes Zhang & Chen, 2015, S. (S.) alafoliacea Zhang & Chen, sp. nov., S. (S.) baoxing Li & Chen, sp. nov., S. (S.) bibarbata Li & Chen, sp. nov., S. (S.) bimai Cui & Chen, sp. nov., S. (S.) cinereipecta Zhang & Chen, sp. nov., S. (S.) cardua Cui & Chen, sp. nov., S. (S.) cordhirsuta Wang & Chen, sp. nov., S. (S.) cornuta Li & Chen, sp. nov., S. (S.) cucullata Li & Chen, sp. nov., S. (S.) cultella Cui & Chen, sp. nov., S. (S.) curvitabulata Cui & Chen, sp. nov., S. (S.) daiya Cui & Chen, sp. nov., S. (S.) dendrophila Zhang & Chen, sp. nov., S. (S.) flabella Li & Chen, sp. nov., S. (S.) flavipes Li & Chen, sp. nov., S. (S.) formosa Zhang & Chen, sp. nov., S. (S.) fusca Li & Chen, sp. nov., S. (S.) fuscipes Li & Chen, sp. nov., S. (S.) glaucopalpula Cui & Chen, sp. nov., S. (S.) haba Zhang & Chen, sp. nov., S. (S.) hirticlavata Cui & Chen, sp. nov., S. (S.) iaspidea Zhang & Chen, sp. nov., S. (S.) idiasta Cui & Chen, sp. nov., S. (S.) kanda Cui & Chen, sp. nov., S. (S.) labao Li & Chen, sp. nov., S. (S.) lancang Li & Chen, sp. nov., S. (S.) latifoliacea Wang & Chen, sp. nov., S. (S.) liusanjieae Li & Chen, sp. nov., S. (S.) magniflava Cui & Chen, sp. nov., S. (S.) mailangang Li & Chen, sp. nov., S. (S.) marenubila Cui & Chen, sp. nov., S. (S.) menghai Zhang & Chen, sp. nov., S. (S.) menglian Li & Chen, sp. nov., S. (S.) minutiflava Li & Chen, sp. nov., S. (S.) multiprocera Li & Chen, sp. nov., S. (S.) nayun Li & Chen, sp. nov., S. (S.) nigridentata Wang & Chen, sp. nov., S. (S.) nigripalpula Cui & Chen, sp. nov., S. (S.) otphylla Cui & Chen, sp. nov., S. (S.) radiciflava Zhang & Chen, sp. nov., S. (S.) rava Cui & Chen, sp. nov., S. (S.) sciophila Li & Chen, sp. nov., S. (S.) septencolorata Li & Chen, sp. nov., S. (S.) serrata Zhang & Chen, sp. nov., S. (S.) silvestrella Zhang & Chen, sp. nov., S. (S.) simola Cui & Chen, sp. nov., S. (S.) yani Li & Chen, sp. nov., S. (S.) yixiang Zhang & Chen, sp. nov., S. (S.) zaduo Cui & Chen, sp. nov., and S. (S.) zhuoma Cui & Chen, sp. nov. We also provided a complete list of Chinese Steganina species together with their geographical distributions. In addition, the majority of currently available DNA barcode (partial sequence of the mitochondrial cytochrome c oxidase subunit I (COI) gene) sequences of this subgenus (435 sequences of 102 spp.) were employed in a molecular analysis for species delimitation. Taken together, morphology- and molecular-based species delimitation results reached a consensus for an overwhelming majority of these Steganina species (98 of 102 spp.).

Effect of Noradrenaline on the Facial Stimulation-Evoked Mossy Fiber-Granule Cell Synaptic Transmission in Mouse Cerebellar Cortex.

Noradrenaline is an important neuromodulator in the cerebellum. We previously found that noradrenaline depressed cerebellar Purkinje cell activity and climbing fiber-Purkinje cell synaptic transmission in vivo in mice. In this study, we investigated the effect of noradrenaline on the facial stimulation-evoked cerebellar cortical mossy fiber-granule cell synaptic transmission in urethane-anesthetized mice. In the presence of a γ-aminobutyrateA (GABAA) receptor antagonist, air-puff stimulation of the ipsilateral whisker pad evoked mossy fiber-granule cell synaptic transmission in the cerebellar granular layer, which expressed stimulus onset response, N1 and stimulus offset response, N2. Cerebellar surface perfusion of 25 µM noradrenaline induced decreases in the amplitude and area under the curve of N1 and N2, accompanied by an increase in the N2/N1 ratio. In the presence of a GABAA receptor blocker, noradrenaline induced a concentration-dependent decrease in the amplitude of N1, with a half-maximal inhibitory concentration of 25.45 µM. The noradrenaline-induced depression of the facial stimulation-evoked mossy fiber-granule cell synaptic transmission was reversed by additional application of an alpha-adrenergic receptor antagonist or an alpha-2 adrenergic receptor antagonist, but not by a beta-adrenergic receptor antagonist or an alpha-1 adrenergic receptor antagonist. Moreover, application of an alpha-2 adrenergic receptor agonist, UK14304, significantly decreased the synaptic response and prevented the noradrenaline-induced depression. Our results indicate that noradrenaline depresses facial stimulation-evoked mossy fiber-granule cell synaptic transmission via the alpha-2 adrenergic receptor in vivo in mice, suggesting that noradrenaline regulates sensory information integration and synaptic transmission in the cerebellar cortical granular layer.

Systematic Investigation of Plant-Parasitic Nematodes Associated with Main Subtropical Crops in Guangxi Province, China.

Plant parasitic nematodes (PPNs) are a pathogenic group that causes momentous crop yield loss by retarding plant growth and development through plant parasitization. In this study, the distribution of PPNs based on the main crops in Guangxi Province of China was investigated. A total of 425 samples of soil or roots from sugarcane, rice, maize, and soybean were collected in 68 counties, and a total of 48 order/family/genera of PPNs were identified, of which some genera were found in more than one crop. A total of 31 order/family/genera of PPNs were found in rice, among which Hirschmanniella was the most abundant, accounting for 79.23%, followed by Tylenchorhynchus (34.43%). Forty order/family/genera were observed in maize, of which the dominant genera were Pratylenchus and Tylenchorhynchus at 45.14% and 32.64%, respectively. In addition, 30 order/family/genera of PPNs were detected from sugarcane, and the percentages of Tylenchorhynchus and Helicotylenchus were 70.42% and 39.44%, respectively. The main crop of Eastern ecological regions was rice, with a high frequency of Hirschmanniella. The greatest frequency of Pratylenchus was found in the Western eco-region, which had a large area of maize. In the Northern eco-region, rice and maize were popular, with abundant Hirschmanniella and Helicotylenchus. In the Central eco-region, Pratylenchus was detected on the main crop of sugarcane. Hirschmanniella (72.94%) was dominant in clay, and Tylenchorhynchus (54.17%) showed the highest frequency in loam. The distribution of PPNs varied with different altitudes. The diversity of this phenomenon was closely related to host plants. These results could improve understanding of the distribution of PPNs and provide important information for controlling PPNs.

Potentially Overestimated Efficacy of Nanoparticle Albumin-bound Paclitaxel compared with Solvent-based Paclitaxel in Breast Cancer: A Systemic Review and Meta-analysis.

Background: Nanoparticle albumin-bound paclitaxel (nab-PTX) has exhibited clinical efficacy in breast cancer treatment, but toxicities can be yielded more at the same time. We did this meta-analysis aiming to unambiguously compare nab-PTX with conventional solvent-based paclitaxel (sb-PTX) in breast cancer patients of all stages. Method: Pubmed, Embase and Cochrane Library were searched for head-to-head randomized controlled trials of nab-PTX and sb-PTX in breast cancer. Risk ratio (RR) with 95% confidence interval was used for dichotomous variables while Hazard ratio (HR) was used for time-to-event outcomes. Results: Our review finally included 9 studies with 3508 patients. Nab-PTX showed a benefit on objective response rate (ORR) (RR=1.22 [1.04-1.43], P=0.01) as well as non-inferiority compared with sb-PTX in disease control rate (DCR) (RR=1.01 [0.98-1.04], P=0.44), overall survival (OS) (HR=0.99 [0.93-1.05], P=0.81) and disease free survival/progression free survival (DFS/PFS) (HR=0.92 [0.81-1.05], P=0.21). However, when it comes to toxicities (fatigue, nausea or vomiting, peripheral sensory neuropathy and adverse event related discontinuation), results favored sb-PTX (RR=2.89 [1.07-7.8], 3.15 [1.78-5.59], 2.11 [1.32-3.37], 2.02 [1.61-2.53]; P<0.05). Patients with metastatic tumors or undergoing conventional schedule responses better to nab-PTX than the compared groups (RR of ORR in metastatic vs early or locally advanced patients: 1.46 [1.09-1.96] vs 1.01 [0.94-1.08]; conventional vs dose dense group: 1.59 [1.23-2.06] vs 1.01 [0.91-1.12]). Conclusions: Nab-PTX can improve ORR compared with paclitaxel and should be given priority to when aiming to reduce tumor load in breast cancer. Sb-PTX of dose dense schedule is recommended when toxicity of nab-PTX is hard to bear for breast cancer patients.

Chronic Ethanol Exposure Enhances Facial Stimulation-Evoked Mossy Fiber-Granule Cell Synaptic Transmission via GluN2A Receptors in the Mouse Cerebellar Cortex.

Sensory information is transferred to the cerebellar cortex via the mossy fiber-granule cell (MF-GC) pathway, which participates in motor coordination and motor learning. We previously reported that chronic ethanol exposure from adolescence facilitated the sensory-evoked molecular layer interneuron-Purkinje cell synaptic transmission in adult mice in vivo. Herein, we investigated the effect of chronic ethanol exposure from adolescence on facial stimulation-evoked MF-GC synaptic transmission in the adult mouse cerebellar cortex using electrophysiological recording techniques and pharmacological methods. Chronic ethanol exposure from adolescence induced an enhancement of facial stimulation-evoked MF-GC synaptic transmission in the cerebellar cortex of adult mice. The application of an N-methyl-D-aspartate receptor (NMDAR) antagonist, D-APV (250 µM), induced stronger depression of facial stimulation-evoked MF-GC synaptic transmission in chronic ethanol-exposed mice compared with that in control mice. Chronic ethanol exposure-induced facilitation of facial stimulation evoked by MF-GC synaptic transmission was abolished by a selective GluN2A antagonist, PEAQX (10 µM), but was unaffected by the application of a selective GluN2B antagonist, TCN-237 (10 µM), or a type 1 metabotropic glutamate receptor blocker, JNJ16259685 (10 µM). These results indicate that chronic ethanol exposure from adolescence enhances facial stimulation-evoked MF-GC synaptic transmission via GluN2A, which suggests that chronic ethanol exposure from adolescence impairs the high-fidelity transmission capability of sensory information in the cerebellar cortex by enhancing the NMDAR-mediated components of MF-GC synaptic transmission in adult mice in vivo.

SANT, a novel Chinese herbal monomer combination, decreasing tumor growth and angiogenesis via modulating autophagy in heparanase overexpressed triple-negative breast cancer.

ETHNOPHARMACOLOGICAL RELEVANCE: Astragalus mongholicus, Solanum nigrum Linn, Lotus plumule, Ligusticum are widely used traditional herbal medicines for cancer treatment in China. They were typical drugs selected from Gubenyiliu II and series of formula (GYII), which were developed on the foundation of YIQIHUOXUEJIEDU theory. In the present study, four active ingredients (Astragaloside IV, α-solanine, neferine, and 2,3,5,6-tetramethylpyrazine) derived from medicines above were applied in combination as SANT. AIM OF THE STUDY: Triple-negative breast cancer (TNBC) is a serious threat to women's health worldwide. Heparanase (HPSE) is often up-regulated in breast cancer with the properties of facilitating tumorigenesis and influencing the autophagy process in cancer cells. This study aimed at evaluating the anti-tumor potential of SANT in treating HPSE related TNBC both in-vitro and in-vivo. MATERIALS AND METHODS: In this study, we explored the correlation between HPSE expression and survival of breast cancer patients in databases. We performed MTS, trans-well and wound scratch assays to assess the impact of SANT on cell proliferation and migration. Confocal microscopy observation and western blots were applied to verify the autophagy flux induced by SANT. Mice models were employed to evaluate the efficacy and safety of SANT in-vivo by tumor weights and volumes or serum index, respectively. To analyze the underlying mechanisms of SANT, we conducted human autophagy PCR array and angiogenesis proteome profiler on tumor tissues. RESULTS: Patients with elevated HPSE expression were associated with a poor outcome in both RFS (P = 1.7e-12) and OS (P = 0.00016). SANT administration significantly inhibited cancer cells' proliferation and migration, enhanced autophagy flux, and slightly reduced the active form of HPSE in-vitro. SANT also suppressed tumor growth and angiogenesis in-vivo. Human autophagy PCR array results indicated that SANT increased the ATG16L1, ATG9B, ATG4D gene expressions while decreased TMEM74 and TNF gene expressions.Angiogenesis proteome profiler results showed SANT reduced protein level of HB-EGF, thrombospondin-2, amphiregulin, leptin, IGFBP-9, EGF, coagulation factor III, and MMP-9 (pro and active form) in tumor, raised the protein expression of serpin E1 and platelet factor 4. CONCLUSIONS: These findings indicated that herbal compounds SANT may be a promising candidate in anti-cancer drug discovery. It also provides novel strategies for using natural compounds to achieve optimized effect.

Erratum: Incidence of the Rice Root-Knot Nematode, Meloidogyne graminicola, in Guangxi, China.

[This corrects the article on p. 297 in vol. 36, PMID: 32547345.].

NMDARs contribute to the facial stimuli-evoked mossy fiber-granule cell synaptic transmission in vivo in mice.

N-methyl-D-aspartate receptors (NMDARs) are expressed in granule cell and involve in mossy fiber-granule cell (MF-GC) synaptic transmission in cerebellar cortex. In the absence GABAA receptor activity, we here studied the role of NMDARs during the facial stimulation evoked MF-GC synaptic transmission in urethane-anesthetized mice using electrophysiological recording technique and pharmacological methods. Our results showed that facial stimuli train (20 Hz, 5 pulses) evoked 5 field potential responses (N1-N5) in mouse cerebellar granular layer, which identified MF-GC synaptic transmission. Blocking NMDARs induced significant depression in the amplitude of N2 to N5, accompanied with significant decrease in pulse ratios, area under the curve (AUC) and half-width of N1. A selective GluN2A antagonist, PEAQX (10 µM) also produced significant depression in the amplitude of N2 to N5, and decreases in pulse ratios. However, a selective GluN2B antagonist, TCN-237 (10 µM) did not significantly attenuate the facial stimuli train-induced mossy fiber-granule cell synaptic transmission. Application of NMDA (1 µM) produced increases in the AUC and half-width of Ron, as well the amplitude and AUC of Roff, which was reversed by following application of PEAQX. Our present results indicated that NMDARs, especially GluN2A contribute to the facial stimulation-evoked MF-GC synaptic transmission, suggesting that the NMDARs play an important role during the lateral sensory information synaptic transmission in the cerebellar granular layer in vivo in mice.

Incidence of the Rice Root-Knot Nematode, Meloidogyne graminicola, in Guangxi, China.

Root-knot nematodes (Meloidogyne spp.) are the most destructive group of plant-parasitic nematodes. Plants infected by Meloidogyne spp. develop above-ground symptoms, stunting, yellowing, nutrient deficiencies, and gall formations with typical hook-shaped root tips. Infected plants experience yield losses. During 2018-2019 survey, leaf chlorosis rice plants were found in 206 fields of 67 counties in Guangxi, China, around 30 days after transplanting. Galls and hooked tips on the roots and pear-shaped females were observed. About 32.04% of fields were infested with the nematode. The nematodes were identified as Meloidogyne graminicola base on morphological and molecular analysis. To the best of our knowledge, this is the first report of M. graminicola on rice plants in Guangxi, China. The results of this study urge the discovery of resistant cultivars and the development of management strategies.

Noradrenaline inhibits complex spikes activity via the presynaptic PKA signaling pathway in mouse cerebellar slices.

Norepinephrine (NA) is an important neurotransmitter of the cerebellum that regulates synaptic transmission, motor regulation and motor learning under certain conditions via adrenergic receptors (ARs). We previously found that NA depressed cerebellar climbing fiber-Purkinje cell (CF-PC) synaptic transmission via α2-ARs in vivo in mice. We here investigated the mechanisms of NA inhibited CF-PC synaptic transmission in acute cerebellar slices using the whole-cell recording technique and pharmacological methods. Bath application of NA (10 µM) depressed CF-PC synaptic transmission, which exhibited a time-dependent decrease in amplitude of excitatory postsynaptic currents (N1), accompanied by an increase in the paired-pulse ratio (PPR). The NA-induced depression of CF-PC synaptic transmission was significantly prevented by inhibition of protein kinase A (PKA) with either H-89 or KT5720. Furthermore, the NA-induced inhibition of CF-PC synaptic transmission was rescued by activation adenylate cyclase (AC), and the AC-induced enhancement of CF-PC synaptic transmission was depressed by NA. Moreover, inhibition of AC with SQ22536, produced a significant depression of CF-PC synaptic transmission and abrogated the NA-induced depression of CF-PC synaptic transmission. However, the NA-induced depression of CF-PC synaptic transmission was not blocked by intracellular inhibition of PKA with a cell impermeable PKA inhibitor, PKI, or by extracellular inhibition of protein kinase C. These results indicate that NA activates presynaptic α2-AR, resulting in a depression of mouse cerebellar CF-PC synaptic transmission through the AC-PKA signaling pathway.

Whole genome sequencing and comparative genomic analysis of oleaginous red yeast Sporobolomyces pararoseus NGR identifies candidate genes for biotechnological potential and ballistospores-shooting.

BACKGROUND: Sporobolomyces pararoseus is regarded as an oleaginous red yeast, which synthesizes numerous valuable compounds with wide industrial usages. This species hold biotechnological interests in biodiesel, food and cosmetics industries. Moreover, the ballistospores-shooting promotes the colonizing of S. pararoseus in most terrestrial and marine ecosystems. However, very little is known about the basic genomic features of S. pararoseus. To assess the biotechnological potential and ballistospores-shooting mechanism of S. pararoseus on genome-scale, the whole genome sequencing was performed by next-generation sequencing technology. RESULTS: Here, we used Illumina Hiseq platform to firstly assemble S. pararoseus genome into 20.9 Mb containing 54 scaffolds and 5963 predicted genes with a N50 length of 2,038,020 bp and GC content of 47.59%. Genome completeness (BUSCO alignment: 95.4%) and RNA-seq analysis (expressed genes: 98.68%) indicated the high-quality features of the current genome. Through the annotation information of the genome, we screened many key genes involved in carotenoids, lipids, carbohydrate metabolism and signal transduction pathways. A phylogenetic assessment suggested that the evolutionary trajectory of the order Sporidiobolales species was evolved from genus Sporobolomyces to Rhodotorula through the mediator Rhodosporidiobolus. Compared to the lacking ballistospores Rhodotorula toruloides and Saccharomyces cerevisiae, we found genes enriched for spore germination and sugar metabolism. These genes might be responsible for the ballistospores-shooting in S. pararoseus NGR. CONCLUSION: These results greatly advance our understanding of S. pararoseus NGR in biotechnological potential and ballistospores-shooting, which help further research of genetic manipulation, metabolic engineering as well as its evolutionary direction.

Chronic ethanol exposure facilitates facial-evoked MLI-PC synaptic transmission via nitric oxide signaling pathway in vivo in mice.

Ethanol (EtOH) exposure causes alterations of motor coordination, balance, behavior, speech, and certain cognitive functions are considered to be caused partly by impairment of cerebellar circuits function and modulation of synaptic transmission. The cerebellar cortical molecular layer interneuron-Purkinje cell (MLI-PC) synapses are critical for various information integration and transmission, which are sensitive to acute and chronic EtOH exposure. The aim of this study is to investigate the effect of chronic ethanol exposure on the facial stimulation-evoked MLI-PC synaptic transmission in urethane-anesthetized mice, by electrophysiological recording and pharmacological methods. Under current-clamp recording conditions, air-puff stimulation of ipsilateral whisker pad evoked MLI-PC synaptic transmission, which expressed an inhibitory component (P1) followed by a pause of simple spike (SS) firing in cerebellar PCs. Chronic ethanol exposure did not change the latency of the facial stimulation-evoked responses in cerebellar PCs, but induced significant enhancement of the stimulation-evoked MLI-PC synaptic transmission, which expressed increases in amplitude of P1 and pause of SS firing. The amplitude of P1 and pause of SS in ethanol exposure group were significant higher than that in control group. Cerebellar surface application of nitric oxide synthesis (NOS) inhibitor, L-NNA (5 mM) significantly decreased the amplitude of P1 and the pause of SS firing in EtOH exposure group, but did no effect on control group. In contrast, cerebellar surface application of NO donor, SNAP (100 µM) significantly increased the amplitude of P1 and the pause of SS firing in control group, but not in EtOH exposure group. These results indicated that chronic EtOH exposure significantly facilitated the sensory-evoked MLI-PC synaptic transmission via NO signaling pathway in mouse cerebellar cortex.

Noradrenaline depresses spontaneous complex spikes activity of cerebellar Purkinje cells via α2-adrenergic receptor in vivo in mice.

Locus coeruleus (LC) noradrenergic neurons afferents release noradrenaline (NA) in the cerebellar cortex for modulating cerebellar neuronal circuitry function. Our previous study found that NA inhibited the spontaneous simple spikes activity of cerebellar Purkinje cells (PC) through activation of molecular layer interneurons (MLIs) in vivo in mice. We here examined the effects of NA on spontaneous complex spikes (CSs) activity of cerebellar PC in urethane-anesthetized mice by electrophysiology recording technique and pharmacological methods. Our results showed that cerebellar surface perfusion of NA significantly reduced the number of spikelets and the area under curve (AUC) of the spontaneous CSs. Application of nonselective adrenergic receptor (AR) antagonist, phentolamine, abolished the NA-induced inhibition of CSs. However applying a nonselective ß-AR blocker, propranolol, failed to prevent the NA-induced inhibition of CSs activity. The NA-induced inhibition of CSs activity was not blocked by α1-AR antagonist, prazosin, but it was abolished by α2-AR antagonist, yohimibine. Moreover, application of α2-AR agonist, UK14304 induced a depression of CSs activity and mimicked the NA-induced inhibition of CS activity. These results indicate that NA regulates spontaneous CSs activity of cerebellar PCs via activation of α2-AR in vivo in mice. Our present results suggest that noradrenergic neurons of LC may modulate the outputs of cerebellar PCs via inhibition of CSs activity.

Estrogen-induced Tgfbr1 and Bmpr1a Expression Repressed via Estrogen Receptor Beta in MC3T3-E1 Cells.

BACKGROUND: Estrogen, as an important hormone in human physiological process, is closely related to bone metabolism. The aim of this study was to investigate the mechanism of estrogen on osteoblasts metabolism in MC3T3-E1 cells. METHODS: We treated the MC3T3-E1 cells with different concentrations of ß-estradiol (0.01, 0.1, 1, and 10 nmol/L), observed the morphological changes of the cells, and detected the cell's proliferation and apoptosis of MC3T3-E1 cells. Two transcriptome libraries were constructed and sequenced. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to confirm the differentially expressed genes (DEGs), and then treated the MC3T3-E1 cells with estrogen receptor (ER) inhibitors α and ß, respectively, and then examined the expression of Tgfbr1 and Bmpr1a genes. The promoter of Tgfbr1 and Bmpr1a gene was analyzed, and the ER response elements were identified. Finally, ChIP was used to verify the binding of ER to Tgfbr1 and Bmpr1a promoter. RESULTS: In the high-concentration ß-estradiol treatment group (1 nmol/L and 10 nmol/L), there was no significant difference in the morphology of the cells under the microscope, 1 nmol/L and 10 nmol/L treated group appeared statistically significant difference in cell apoptosis and proliferation (P < 0.05 and P < 0.01, respectively). We found 460 DEGs compared with the control group. Among the DEGs, there were 66 upregulated genes and 394 downregulated genes. Gene ontology classification and Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed that many bone metabolism-related biological processes and cell signaling pathways were disordered. The qRT-PCR verification showed that the expressions of Tgfbr1- and Bmpr1a-related genes in bone metabolism pathway in the 10 nmol/L treatment group were significantly decreased (P < 0.05). ER ß was involved in the inhibitory effect of Tgfbr1 and Bmpr1a genes. The bioinformatics of the promoter found that there were three ER response elements in the promoter of Tgfbr1, and there were two ER response elements in Bmpr1a promoter regions. ChIP experiments showed that estrogen could enhance the binding of ERs to Tgfbr1 and Bmpr1a genes. CONCLUSIONS: Estrogen can promote the apoptosis and proliferation of osteoblasts simultaneously, and the mechanism may be the joint action of transforming growth factor-beta, Wnt, mitogen-activated protein kinase, and nuclear factor-kappaB bone metabolism-related signaling pathway. Estrogen inhibits the expression of Tgfbr1 and Bmpr1a genes through ER ß and affects the metabolism of MC3T3-E1 osteoblasts.

Corticotrophin-Releasing Factor Modulates Cerebellar Purkinje Cells Simple Spike Activity in Vivo in Mice.

Corticotropin-releasing factor (CRF) is a major neuromodulator that modulates cerebellar neuronal activity via CRF receptors during stress responses. In the cerebellar cortex, CRF dose-dependently increases the simple spike (SS) firing rate of Purkinje cells (PCs), while the synaptic mechanisms of this are still unclear. We here investigated the effect of CRF on the spontaneous SS activity of cerebellar PCs in urethane-anesthetized mice by in vivo electrophysiological recording and pharmacological methods. Cell-attached recordings from PCs showed that micro-application of CRF in cerebellar cortical molecular layer induced a dose-dependent increase in SS firing rate in the absence of GABAA receptor activity. The CRF-induced increase in SS firing rate was completely blocked by a nonselective CRF receptor antagonist, α-helical CRF-(9-14). Nevertheless, application of either a selective CRF-R1 antagonist, BMS-763534 (BMS, 200 nM) or a selective CRF-R2 antagonist, antisauvagine-30 (200 nM) significantly attenuated, but failed to abolished the CRF-induced increase in PCs SS firing rate. In vivo whole-cell patch-clamp recordings from PCs showed that molecular layer application of CRF significantly increased the frequency, but not amplitude, of miniature postsynaptic currents (mEPSCs). The CRF-induced increase in the frequency of mEPSCs was abolished by a CRF-R2 antagonist, as well as protein kinase A (PKA) inhibitors. These results suggested that CRF acted on presynaptic CRF-R2 of cerebellar PCs resulting in an increase of glutamate release through PKA signaling pathway, which contributed to modulation of the cerebellar PCs outputs in Vivo in mice.

[Ursolic acid activates chloride channels and decreases cell volume in poorly differentiated nasopharyngeal carcinoma cells].

The present study aimed to investigate the effects of ursolic acid on the chloride channels and cell volume in nasopharyngeal carcinoma cells (CNE-2Z). The whole-cell patch clamp technique was used to detect the current, and cell imaging technique was applied to measure cell volume. The properties of the currents induced by ursolic acid were investigated by changing the extracellular osmotic pressure, replacing the extracellular anions and applying chloride channel blockers. The results showed that, under isotonic conditions, the background current was weak and stable. When perfusing the cells with ursolic acid (100 nmol/L), a large current (-59.86 pA/pF ± 4.86 pA/pF at -80 mV, 78.92 pA/pF ± 6.39 pA/pF at +80 mV) was induced. The chloride current showed outward rectification and negligible time- and voltage-dependent inactivation. The reversal potential (-4.83 mV ± 0.30 mV) of the current was close to the calculated equilibrium potential for Cl⁻ (-0.9 mV). The permeabilities of the channel to different anions were ranked in order as follows: Cl⁻ = I⁻ > Br⁻ > gluconate. Hypertonic solutions inhibited the current induced by ursolic acid. The chloride channel blockers, tamoxifen (20 µmol/L) and 5-nitro-2-(3-phenylpro-pylamino) benzoic acid (NPPB, 100 µmol/L), suppressed the current. Furthermore, ursolic acid decreased the cell volume by (11.78 ± 1.20)% in 1 h, and the effect was inhibited by NPPB. These results suggest that ursolic acid can activate chloride channels, resulting in outflow of Cl⁻ and decrease of cell volume in nasopharyngeal carcinoma cells.

[Clinical significance of detecting partial cytokines in serum of patients with pulmonary tuberculosis by cytometric bead array].

AIM: To explore the clinical significance of cytometric bead array(CBA) in detecting partial cytokines(IL-2, IL-4, IL-6, IL-10, TNF-α and IFN-γ)in the serum from patients with different types of pulmonary tuberculosis. METHODS: The levels of six cytokines (IL-2, IL-4, IL-6, IL-10, TNF-α and IFN-γ) in the serum from 84 cases of patients with pulmonary tuberculosis(46 cases of active pulmonary tuberculosis patients, 38 cases of inactive pulmonary tuberculosis patients) and 30 cases of normal person, were detected by BD CBA Flex Set kit. RESULTS: The levels of IL-2, IL-4, IL-6, IL-10, TNF-α and IFN-γ in pulmonary tuberculosis group were significantly higher than those in control group (P<0.01 or P<0.05.Compared with the inactive pulmonary tuberculosis group, the levels of serum IL-2, IL-4, IL-6, IL-10, TNF-α and IFN-γ in the active pulmonary tuberculosis group were significantly higher (P<0.01 or P<0.05).Serum IL-4 and TNF-α levels in active pulmonary tuberculosis group, inactive pulmonary tuberculosis group and the control group had not significantly different. CONCLUSION: Six cytokines in serum can be detected quickly and sensitively in batches at one time by CBA. IL-2, IL-4, IL-6, IL-10, TNF-α and IFN-γ play an important role in the immune pathogenesis of pulmonary tuberculosis.

Optimized medium improves expression and secretion of extremely thermostable bacterial xylanase, XynB, in Kluyveromyces lactis.

An extremely thermostable xylanase gene, xynB, from hyperthermophilic bacterium Thermotoga maritima MSB8 was successful expressed in Kluyveromyces lactis. Response surface methodology (RSM) was applied to optimize medium components for production of XynB secreted by the recombinant K. lactis. Secretion level (102 mg/L) and enzyme activity (49 U/ml) of XynB in the optimized medium (yeast extract, lactose, and urea; YLU) were much higher than those (56 mg/L, 16 U/ml) in original medium (yeast extract, lactose, and peptone; YLP). It was also observed that the secretory efficiency of mature XynB was improved by the YLU medium. mRNA levels of 13 characterized secretion-related genes between K. lactis cultured in YLP and YLU were detected using semi-quantitative RT-PCR method. It was found that unfolded protein response (UPR) related genes such as ero1, hac1, and kar2 were up-regulated in K. lactis cultured in YLU. Therefore, nutrient ingredient, especially nitrogen source had a significant influence on the XynB secretory efficiency in the host K. lactis.

Production of pigment-free pullulan by swollen cell in Aureobasidium pullulans NG which cell differentiation was affected by pH and nutrition.

A black yeast strain "NG" was isolated from strawberry fruit and identified as Aureobasidium pullulans. Strain NG displayed yeast-like cell (YL), swollen cell (SC), septate swollen cell (SSC), meristematic structure (MS), and chlamydospore (CH) morphologies. pH was the key factor regulating cell morphogenesis of strain NG. Differentiation of YL controlled by extracellular pH had no relationship with nutrition level. YL was maintained at pH >6.0, but was transformed into SC at pH approximately 4.5. SC, a stable cell type of A. pullulans, could bud, septate, or transform into MS or CH, in response to nutrition level and low pH. SC produced swollen cell blastospores (SCB) at pH 2.1 with abundant nutrition, and could transform into MS at lower pH (1.5). SC was induced to form CH by low level nutrition and pH <3, and this transition was suppressed by adjusting pH to approximately 4.5. Crude polysaccharides without pigment (melanin) were produced by SC of strain NG. Pullulan content of the polysaccharides was very high (98.37%). Fourier-transform infrared spectroscopy confirmed that chemical structures of the polysaccharides and standard pullulan were identical. Swollen cells produced 2.08 mg/ml non-pigmented polysaccharides at 96 h in YPD medium. Controlling pH of fermentation is an effective and convenient method to harvest SC for melanin-free pullulan production.