Ovomucin Ameliorates Intestinal Barrier and Intestinal Bacteria to Attenuate DSS-Induced Colitis in Mice.

Egg white ovomucin (OVM) is homologically related to MUC2, the key component of colonic mucous layer. This study investigated the effects of orally administered OVM from egg white on the colonic mucosal barrier and the development of colitis using a colitis C57BL/6J mice model. The results showed that daily supplementation of 125 and 250 mg/kg BW of OVM partially relieved the villous destruction and loss of intestinal barrier integrity, and hence decreased the epithelial barrier permeability. The supplementation also reduced the secretion of proinflammatory cytokines TNF-α and IL-6. Besides, OVM administration significantly increased the relative abundance of intestinal beneficial bacteria including Lactobacilli, Faecalibaculum, Ruminococcus, etc. and further upregulated the production of bacterial metabolites such as short-chain fatty acids (SCFAs), which is a direct source of energy for the proliferation of epithelia and goblet cells. In conclusion, OVM from egg white ameliorates colitis by enhancing the intestinal barrier function and abundance of intestinal bacteria, thereby increasing the number of SCFAs.

Integrated analysis of gene expression and copy number variations in MET proto­oncogene­transformed human primary osteoblasts.

The aim of the present study was to screen the potential osteosarcoma (OS)­associated genes and to obtain additional insight into the pathogenesis of OS. Transcriptional profile (ID: GSE28256) and copy number variations (CNV) profile were downloaded from Gene Expression Omnibus database. Differentially expressed genes (DEGs) between MET proto­oncogene­transformed human primary osteoblast (MET­HOB) samples and the control samples were identified using the Linear Models for Microarray Data package. Subsequently, CNV areas and CNVs were identified using cut­off criterion of >30%­overlap within the cases using detect_cnv.pl in PennCNV. Genes shared in DEGs and CNVs were obtained and discussed. Additionally, the Database for Annotation, Visualization and Integrated Discovery was used to identify significant Gene Ontology (GO) functions and pathways in DEGs with P<0.05. A total of 1,601 DEGs were screened out in MET­HOBs and compared with control samples, including 784 upregulated genes, such as E2F transcription factor 1 (E2F1) and 2 (E2F2) and 817 downregulated genes, such as retinoblastoma 1 (RB1) and cyclin D1 (CCND1). DEGs were enriched in 344 GO terms, such as extracellular region part and extracellular matrix and 14 pathways, including pathways in cancer and extracellular matrix­receptor interaction. Additionally, 239 duplications and 439 deletions in 678 genes from 1,313 chromosome regions were detected. A total of 12 genes were identified to be CNV­driven genes, including cadherin 18, laminin subunit α 1, spectrin ß, erythrocytic, ciliary rootlet coiled­coil, rootletin pseudogene 2, ß­1,4-N-acetyl-galactosaminyltransferase 1, G protein regulated inducer of neurite outgrowth 1, EH domain binding protein 1­like 1, growth factor independent 1, cathepsin Z, WNK lysine deficient protein kinase 1, glutathione S­transferase mu 2 and microsomal glutathione S­transferase 1. Therefore, cell cycle­associated genes including E2F1, E2F2, RB1 and CCND1, and cell adhesion­associated genes, such as CDH18 and LAMA1 may be used as diagnosis and/or therapeutic markers for patients with OS.

Α-Dendrotoxin-sensitive Kv1 channels contribute to conduction failure of polymodal nociceptive C-fibers from rat coccygeal nerve.

It is known that some patients with diabetic neuropathy are usually accompanied by abnormal painful sensations. Evidence has accumulated that diabetic neuropathic pain is associated with the hyperexcitability of peripheral nociceptors. Previously, we demonstrated that reduced conduction failure of polymodal nociceptive C-fibers and enhanced voltage-dependent sodium currents of small dorsal root ganglion (DRG) neurons contribute to diabetic hyperalgesia. To further investigate whether and how potassium channels are involved in the conduction failure, α-dendrotoxin (α-DTX), a selective blocker of the low-threshold sustained Kv1 channel, was chosen to examine its functional capability in modulating the conduction properties of polymodal nociceptive C-fibers and the excitability of sensory neurons. We found that α-DTX reduced the conduction failure of C-fibers from coccygeal nerve in vivo accompanied by an increased initial conduction velocity but a decreased activity-dependent slowing of conduction velocity. In addition, the number of APs evoked by step currents was significantly enhanced after the treatment with α-DTX in small-diameter sensory neurons. Further study of the mechanism indicates α-DTX-sensitive K(+) current significantly reduced and the activation of this current in peak and steady state shifted to depolarization for diabetic neurons. Expression of Kv channel subunits Kv1.2 and Kv1.6 was downregulated in both small dorsal root ganglion neurons and peripheral C-fibers. Taken together, these results suggest that α-DTX-sensitive Kv1 channels might play an important role in regulating the conduction properties of polymodal nociceptive C-fibers and firing properties of sensory neurons.

[Leptin promotes the expression of hTERT via STAT3 in breast cancer cells].

OBJECTIVE: To discussion the in vitro molecular mechanism of leptin promoting the expression of hTERT in breast cancer cells. METHODS: The hTERT mRNA expression of STAT3 knockdown on leptin-induced hTERT was measured by reverse transcription-polymerase chain reaction (RT-PCR). Determine the expression of hTERT protein after different treatments in MCF7 by Western blot. Chromatin immunoprecipitation assay (ChIP) was performed to detect the binding of STAT3 to hTERT promoter in MCF7. Luciferase assay was used to confirm the effects of leptin and STAT3 phosphorylation inhibitor on the transcriptional activity of hTERT promoter. RESULTS: The RT-PCR analysis showed that knockdown of STAT3 significantly reduced the leptin-induced transcription of hTERT. Western blot showed that the expression of hTERT were 3.109 ± 0.051 and 1.025 ± 0.031 after leptin or both of leptin and AG490 treatments. The results of CHIP showed that the mRNA of control and leptin (160 ng/ml) treatment were 1 and 3.311 ± 0.017. Leptin increased the combination of STAT3 and hTERT promoter. Luciferase assay showed that when the concentration of leptin was 160 ng/ml, the hTERT promoter activity was 80.98 ± 0.18 while the control was 20.76 ± 0.31. After AG490 treatment, the hTERT promoter activity was 18.65 ± 0.32,significantly reduced the leptin-induced activity of hTERT promoter. CONCLUSION: Leptin/STAT3 signaling is a novel pathway for the up-regulation of hTERT expression in breast cancer cells.

Gastrodin inhibits allodynia and hyperalgesia in painful diabetic neuropathy rats by decreasing excitability of nociceptive primary sensory neurons.

Painful diabetic neuropathy (PDN) is a common complication of diabetes mellitus and adversely affects the patients' quality of life. Evidence has accumulated that PDN is associated with hyperexcitability of peripheral nociceptive primary sensory neurons. However, the precise cellular mechanism underlying PDN remains elusive. This may result in the lacking of effective therapies for the treatment of PDN. The phenolic glucoside, gastrodin, which is a main constituent of the Chinese herbal medicine Gastrodia elata Blume, has been widely used as an anticonvulsant, sedative, and analgesic since ancient times. However, the cellular mechanisms underlying its analgesic actions are not well understood. By utilizing a combination of behavioral surveys and electrophysiological recordings, the present study investigated the role of gastrodin in an experimental rat model of STZ-induced PDN and to further explore the underlying cellular mechanisms. Intraperitoneal administration of gastrodin effectively attenuated both the mechanical allodynia and thermal hyperalgesia induced by STZ injection. Whole-cell patch clamp recordings were obtained from nociceptive, capsaicin-sensitive small diameter neurons of the intact dorsal root ganglion (DRG). Recordings from diabetic rats revealed that the abnormal hyperexcitability of neurons was greatly abolished by application of GAS. To determine which currents were involved in the antinociceptive action of gastrodin, we examined the effects of gastrodin on transient sodium currents (I(NaT)) and potassium currents in diabetic small DRG neurons. Diabetes caused a prominent enhancement of I(NaT) and a decrease of potassium currents, especially slowly inactivating potassium currents (I(AS)); these effects were completely reversed by GAS in a dose-dependent manner. Furthermore, changes in activation and inactivation kinetics of I(NaT) and total potassium current as well as I(AS) currents induced by STZ were normalized by GAS. This study provides a clear cellular basis for the peripheral analgesic action of gastrodin for the treatment of chronic pain, including PDN.

Reduced conduction failure of the main axon of polymodal nociceptive C-fibres contributes to painful diabetic neuropathy in rats.

Painful diabetic neuropathy is a common complication of diabetes mellitus and can affect many aspects of life and severely limit patients' daily functions. Signals of painful diabetic neuropathy are believed to originate in the peripheral nervous system. However, its peripheral mechanism of hyperalgesia has remained elusive. Numerous studies have accumulated that polymodal nociceptive C-fibres play a crucial role in the generation and conduction of pain signals and sensitization of which following injury or inflammation leads to marked hyperalgesia. Traditionally, the number of nociceptive primary afferent firings is believed to be determined at the free nerve endings, while the extended main axon of unmyelinated C-fibres only involves the reliable and faithful propagation of firing series to the central terminals. We challenged this classic view by showing that conduction of action potential can fail to occur in response to repetitive activity when they travel down the main axon of polymodal nociceptive C-fibres. Quantitative analysis of conduction failure revealed that the degree of conduction failure displays a frequency-dependent manner. Local administration of low threshold, rapidly activating potassium current blocker, α-dendrotoxin (0.5 nM) and persistent sodium current blocker, low doses of tetrodotoxin (<100 nM) on the main axon of C-fibres can reciprocally regulate the degree of conduction failure, confirming that conduction failure did occur along the main axon of polymodal nociceptive C-fibres. Following streptozotocin-induced diabetes, a subset of polymodal nociceptive C-fibres exhibited high-firing-frequency to suprathreshold mechanical stimulation, which account for about one-third of the whole population of polymodal nociceptive C-fibres tested. These high-firing-frequency polymodal nociceptive C-fibres in rats with diabetes displayed a marked reduction of conduction failure. Delivery of low concentrations of tetrodotoxin and Nav1.8 selective blocker, A-803467 on the main axon of C-fibres was found to markedly enhance the conduction failure in a dose-dependent manner in diabetic rats. Upregulated expression of sodium channel subunits Nav1.7 and Nav1.8 in both small dorsal root ganglion neurons and peripheral C-fibres as well as enhanced transient and persistent sodium current and increased excitability in small dorsal root ganglion neurons from diabetic rats might underlie the reduced conduction failure in the diabetic high-firing-frequency polymodal nociceptive C-fibres. This study shed new light on the functional capability in the pain signals processing for the main axon of polymodal nociceptive C-fibres and revealed a novel mechanism underlying diabetic hyperalgesia.

[Apoptosis resistance induced by leptin and its mechanism in breast cancer cells].

OBJECTIVE: To explore the apoptosis resistance induced by Leptin and its mechanism in breast cancer cells in vitro. METHODS: The leptin-mediated reduction of docetaxel-induced apoptosis in human breast cancer T47D cells was evaluated by TransAM ELISA, MTT and caspase-9 assay. The leptin-promoted survivin expression was analyzed by Western-blot and RT-PCR. The reversing effect of STAT3 knockdown on leptin-induced survivin upregulation was measured by Western-blot and RT-PCR. RESULTS: Leptin promoted T47D cells proliferation and the inhibitory rate was -63.6%. It reduced docetaxel-induced apoptosis in T47D cells by 31.9%. Leptin at different concentrations promoted survivin protein and mRNA expression in T47D cells. The expression of survivin mRNA was 4.6 fold compared with the T47D cells not treated with leptin(10 nmol/L). The expression of survivin mRNA in T47D cells was 0.55 +/- 0.15 fold after transfected with small interfering RNA (siRNA) of STAT3. The expression of survivin mRNA in STAT3 siRNA group and mock transfected group were 0.56 +/- 0.18 fold and 1.61 +/- 0.22 fold after treated by leptin, respectively. The survivin protein level of T47D mock transfected cells was increased after treated by leptin, but the protein level of T47D transfected with STAT3 siRNA cells were not changed significantly. CONCLUSION: Leptin/STAT3 signaling is a novel pathway for up-regulation of survivin expression in breast cancer cells.