Standardized Centella asiatica extract ECa 233 alleviates pain hypersensitivity by modulating P2X3 in trigeminal neuropathic pain

Abstract During oral surgery and temporomandibular joint repositioning, pain hypersensitivity often occurs due to irritation or inflammation of the nerve endings in the orofacial region. Objective: This study aimed to investigate the effects of ECa 233, a Centella asiatica-standardized extract, on the development of mechanical hyperalgesia and allodynia induced by chronic constriction injury of the infraorbital nerve in mice. Methodology: The right infraorbital nerves of the mice were ligated. Oral carbamazepine (20 mg/kg) or ECa 233 (30, 100, or 300 mg/kg) was administered daily for 21 days. Von Frey and air-puff tests were performed on both sides of the whisker pad on days 0, 7, 14, and 21. Thereafter, the expression of purinergic receptor subtype 3 (P2X3) and voltage-gated sodium channel 1.7 (NaV1.7), a transmembrane protein, in the trigeminal ganglion and c-fos immunoreactivity-positive neurons in the trigeminal nucleus caudalis was assessed. Results: After 21 days of infraorbital nerve ligation, the mice showed allodynia- and hyperalgesia-like behavior, P2X3 and NaV1.7 were upregulated in the trigeminal ganglion, and nociceptive activity increased in the trigeminal nucleus caudalis. However, the oral administration of carbamazepine (20 mg/kg), ECa 233 (100 mg/kg), or ECa 233 (300 mg/kg) mitigated these effects. Nevertheless, ECa 233 failed to affect NaV1.7 protein expression. Conclusion: Carbamazepine and ECa 233 can prevent pain hypersensitivity in mice. Considering the side effects of the long-term use of carbamazepine, ECa 233 monotherapy or combined ECa 233 and carbamazepine therapy can be used as an alternative for regulating the development of hypersensitivity in trigeminal pain. However, further detailed clinical studies should be conducted to provide comprehensive information on the use of ECa 233.

Inhibition of subicular seizure-labeled c-fos+ neurons alleviates cognitive deficit in epilepsy / 中国药理学与毒理学杂志

OBJECTIVE Cognitive deficit is a com-mon comorbidity in temporal lobe epilepsy(TLE)and that is not well controlled by current therapeutics.Currently,how epileptic seizure affects cognitive performance remains largely unclear.The subiculum is the major out-put of the hippocampus,which projects to entorhinal cor-tex and other more distinct brain regions.Physiologically,the subiculum codes spatial working memory and naviga-tion information including place,speed,and trajectory.Importantly,prior studies have noted the importance of the subiculum in the beginning,spreading,and generaliz-ing process of hippocampal seizure.How seizure-activated neurons in subiculum participate in cognitive impairment remains largely elusive.METHODS In this study,we sought to label the subicular seizure-activated c-fos+ neu-rons with a special promoter with enhanced synaptic activity-responsive element E-SARE in the subiculum,combined with chemogenetics and designer receptors exclusively activated by designer drugs(DREADDs),Ca2+ fiber photometry approaches,and behavioral tasks,to reveal the role of these neurons in cognitive impairment in epilepsy.RESULTS We found that chemogenetic inhibi-tion of subicular seizure-tagged c-fos+ neurons(mainly CaMK Ⅱ α+ glutamatergic neurons)alleviates seizure generalization and improves cognitive performance in the hippocampal CA3 kindling TLE model.While inhibition of seizure-labeled c-fos+ GABAergic interneuron shows no effect on seizure and cognition.As a comparison,che-mogenetic inhibition of the whole subicular CaMK Ⅱ α+ neuron impairs cognitive function in na?ve mice in basal condition.Notably,inhibition of subicular seizure-tagged c-fos+ neurons enhances the recruitment of cognition-responsive c-fos+ neurons via increasing neural excitability during cognition tasks.CONCLUSION Our results dem-onstrate that subicular seizure-activated c-fos+ neurons contribute to cognitive impairment in TLE,suggesting sei-zure-tagged c-fos+ neurons as the potential therapeutic target to alleviate cognitive impairment in TLE.

Cloning, identification and functional analysis of the goat transcription factor c-fos / 生物工程学报

C-fos is a transcription factor that plays an important role in cell proliferation, differentiation and tumor formation. The aim of this study was to clone the goat c-fos gene, clarify its biological characteristics, and further reveal its regulatory role in the differentiation of goat subcutaneous adipocytes. We cloned the c-fos gene from subcutaneous adipose tissue of Jianzhou big-eared goats by reverse transcription-polymerase chain reaction (RT-PCR) and analyzed its biological characteristics. Using real-time quantitative PCR (qPCR), we detected the expression of c-fos gene in the heart, liver, spleen, lung, kidney, subcutaneous fat, longissimus dorsi and subcutaneous adipocytes of goat upon induced differentiation for 0 h to 120 h. The goat overexpression vector pEGFP-c-fos was constructed and transfected into the subcutaneous preadipocytes for induced differentiation. The morphological changes of lipid droplet accumulation were observed by oil red O staining and bodipy staining. Furthermore, qPCR was used to test the relative mRNA level of the c-fos overexpression on adipogenic differentiation marker genes. The results showed that the cloned goat c-fos gene was 1 477 bp in length, in which the coding sequence was 1 143 bp, encoding a protein of 380 amino acids. Protein structure analysis showed that goat FOS protein has a basic leucine zipper structure, and subcellular localization prediction suggested that it was mainly distributed in the nucleus. The relative expression level of c-fos was higher in the subcutaneous adipose tissue of goats (P < 0.05), and the expression level of c-fos was significantly increased upon induced differentiation of subcutaneous preadipocyte for 48 h (P < 0.01). Overexpression of c-fos significantly inhibited the lipid droplets formation in goat subcutaneous adipocytes, significantly decreased the relative expression levels of the AP2 and C/EBPβ lipogenic marker genes (P < 0.01). Moreover, AP2 and C/EBPβ promoter are predicted to have multiple binding sites. In conclusion, the results indicated that c-fos gene was a negative regulatory factor of subcutaneous adipocyte differentiation in goats, and it might regulate the expression of AP2 and C/EBPβ gene expression.

Roles of c-Fos, EGR-1, PKA, and PKC in cognitive dysfunction in rats after propofol anesthesia

Abstract This study aimed to investigate possible changes in the spatial memory of rats and the expression or activity of EGR-1, c-Fos, PKA, and PKC after propofol anesthesia. Thirty-six Sprague-Dawley rats aged 20 months and 36 Sprague-Dawley rats aged three months were each randomly divided into three groups: the control group, the Morris Water Maze (MWM) group, and the propofol group. In the propofol groups of both young and aged rats, the rats were anesthetized by propofol for two or four hours and then performed the MWM test two days or two weeks after anesthesia to assess cognitive function. EGR-1, c-Fos, PKA, and PKC expressions in the rat hippocampus were determined via immunohistochemistry. For the older rats, the escape latency in the P4h/2d group was significantly prolonged (P < 0.05), and the learning curve was right-shifted in the P4h/2w group (P < 0.05). The expression levels of EGR-1, c-Fos, PKA, and PKC in the MWM groups were significantly higher than those in the control groups (P < 0.05). In the P4h/2d group of aged rats, the expression levels of both PKA and PKC were decreased compared with those of the MWM groups. The decreased expression of both protein kinases may be responsible for the observed impairment after propofol anesthesia

C-Fos directly interacts with mitogen activated protein kinase 14 to regulate the proliferation and apoptosis of rheumatoid arthritis fibroblast-like synovial cells / 中华风湿病学杂志

Objective:To explore the interaction between C-Fos and mitogen activated protein kinase 14 (MAPK14) in rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS), and its effect on the proliferation and apoptosis of RA-FLSs.Methods:RA-FLS and normal fibroblast-like synovial cells (FLS) were cultured. Real-time fluorescent quantitative polymerase chain reaction (RT-qPCR) and Western blotting were used to detect the expression levels of C-Fos mRNA and protein in the two groups. RA-FLS cells were divided into C-Fos overexpression group (transfected with pcDNA3.1-Myc-C-Fos plasmid), overexpression control group (transfected with pcDNA3.1-Myc empty plasmid), and C-Fos silent group (transfection siRNA-C-Fos), silence control group (transfection siRNA-NC) and blank control group (without any treatment). CCK-8 method was used to detect cell proliferation in each group, and flow cytometry was used to detect cell apoptosis in each group. Western blotting was used to detect the expression levels of C-Fos, MAPK14, p-MAPK14, ki-67 and Bax protein in each group. The indirect immunofluorescence experiment analyzed the spatial co-localization of C-Fos and MAPK14, and the co-immunoprecipitation experiment analyzed whether there was interactions between C-Fos and MAPK14 protein. The results of the experimental data were analyzed by Graph Pad Prism 5.0 software. The data of normal distribution was shown as Mean ± standard deviation, and the comparison between the two independent samples using the t test. One-way Analysis of Variance (ANOVA) was used for overall comparison among the multiple groups in the experimental group, and LSD- t test was used for pair comparison within the group. P<0.05 indicated that the difference was statistically significant. Results:The mRNA levels of C-Fos (5.37±0.91) in RA-FLS were significantly higher than FLS (1.46±0.32) ( t=9.94, P<0.001). The protein levels of C-Fos (1.12±0.15) were significantly higher than FLS (0.81±0.07) ( t=3.18, P=0.017). Compared with the blank control group and the overexpression control group, RA-FLS cells transfected with pcDNA3.1-Myc-C-Fos could promote the proliferation of RA-FLS cells, inhibit apoptosis, significantly up-regulate the expression levels of C-Fos, p-MAPK14, ki-67, and significantly down-regulate cellular Bax protein levels (all P<0.05). Compared with the blank control group and the silent control group, RA-FLS cells transfected with siRNA-C-Fos could inhibit the proliferation of RA-FLS cells, promote apoptosis, down-regulate the expression levels of C-Fos, p-MAPK1, ki-67, and up-regulate the cellular Bax protein expression level (all P<0.05). The results of indirect immunofluorescence experiments showed that both C-Fos and MAPK14 could be expressed in the nucleus of RA-FLS. The co-immunoprecipitation experiment verified that C-Fos and MAPK14 protein interact with each other. Conclusion:The interaction of C-Fos-MAPK14 promotes the autophosphorylation of MAPK14, thereby promoting the proliferation of rheumatoid arthritis fibroblast-like synovial cells and inhibiting apoptosis.

Paraventricular Nucleus P2X7 Receptors Aggravate Acute Myocardial Infarction Injury via ROS-Induced Vasopressin-V1b Activation in Rats / 神经科学通报·英文版

The present study was designed to investigate the mechanisms by which P2X7 receptors (P2X7Rs) mediate the activation of vasopressinergic neurons thereby increasing sympathetic hyperactivity in the paraventricular nucleus (PVN) of the hypothalamus of rats with acute myocardial ischemia (AMI). The left anterior descending branch of the coronary artery was ligated to induce AMI in rats. The rats were pretreated with BBG (brilliant blue G, a P2X7R antagonist), nelivaptan (a vasopressin V1b receptor antagonist), or diphenyleneiodonium (DPI) [an nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor]. Hemodynamic parameters of the heart were monitored. Myocardial injury and cardiomyocyte apoptosis were assessed. In the PVN of AMI rats, P2X7R mediated microglial activation, while reactive oxygen species (ROS) and NADPH oxidase 2 (NOX2) were higher than in the sham group. Intraperitoneal injection of BBG effectively reduced ROS production and vasopressin expression in the PVN of AMI rats. Moreover, both BBG and DPI pretreatment effectively reduced sympathetic hyperactivity and ameliorated AMI injury, as represented by reduced inflammation and apoptosis of cardiomyocytes. Furthermore, microinjection of nelivaptan into the PVN improved cardiac function and reduced the norepinephrine (AE) levels in AMI rats. Collectively, the results suggest that, within the PVN of AMI rats, P2X7R upregulation mediates microglial activation and the overproduction of ROS, which in turn activates vasopressinergic neuron-V1b receptors and sympathetic hyperactivity, hence aggravating myocardial injury in the AMI setting.

Analgesic effect of gabapentin combined with agmatine on neuropathic pain in diabetic rats / 国际药学研究杂志

Objective: To investigate analgesic effect of gabapentin(GBP)combined with agmatine(AGM)on diabetic neuropathic pain(DNP)model rats and explore possible mechanism. Methods: SPF SD male rats were injected intraperitoneally with STZ 65 mg/kg to create a neuropathic pain model of diabetic rats. The model rats were randomly divided into 5 groups(n=8): the model group, low-dose GBP group(30 mg/kg, ip), high-dose GBP group(100 mg/kg, ip), AGM group(80 mg/kg, ig)and the GBP-AGM combined group(GBP 30 mg/kg, ip+AGM 80 mg/kg, ig). In addition, a control group was set with 8 randomly selected normal rats. The control group and the model group were intragastrically and intraperitoneally administered an equal volume of physiological saline, respectively, while the test groups were administered drugs with the given dose in the indicated manner, all for continuous 14 days. The rat body mass, tail vein blood glucose, mechanical withdrawal threshold(MWT)and thermal withdrawal latency(TWL) were measured on day 1 before STZ injection and every 7th day after STZ injection, and the plantar tenderness meter was used for the MWT and TWL measurement. The rats were sacrificed 24 h after the last administration, and the spinal cord tissues were harvested. Western blotting was used to detect the expression of p-ERK and c-Fos protein in spinal cord tissues. Results: Compared with the normal control group, the body mass was reduced, blood glucose increased, MWT decreased, and TWL shortened in the model group, all significantly(P0.05)in all of the drug-test groups, while the MWT was increased and the TWL was prolonged in the GBP 100 mg/kg group and the GBP-AGM combined group(both P<0.01). Western blotting results showed that the level of p-ERK and c-Fos protein in the spindal cord was significantly higher in the model group than in the control group(P<0.05). Further, the p-ERK and c-Fos protein level was significantly lower in the GBP+AGM combined group than in the model group(P<0.05)and there was no statistical difference between the GBP 100 mg/kg group and the GBP-AGM combination group. Conclusion: The combination of GBP 30 mg/kg with AGM 80 mg/kg could alleviate neuropathic pain in diabetic rats, which is similar to GBP 100mg/kg and the analgesic effect is likely related to the inhibition of ERK/c-Fos signaling pathway in the spina cord.

Effect of Anemarrhena Asphodeloside BⅡ on Osteoclast Differentiation Based on Molecular Docking / 中国实验方剂学杂志

Objective:To explore the effect of anemarrhena asphodeloside BⅡ (TBⅡ) on the expressions of nuclear transcription factor-κB receptor activator factor ligand （RANKL）, RANK and C-FOS genes during osteoclast differentiation. Method:Molecular docking software LeDock was used to score the docking of TBⅡ with RANKL, RANK and C-FOS. RAW264.7 was treated with soluble RANKL（sRANKL） and divided into control group, sRANKL group (model group), Icariin (Ica) group, low-dose TBIⅡ group (2 μmol·L-1), medium-dose TBⅡ group (4 μmol·L-1), and high-dose TBⅡ group (8 μmol·L-1). The corresponding kit was used to detect iconic enzyme (TRAP) of osteoclast differentiation. Total RNA was extracted by trizol method, Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR) was used to detect the expressions of C-FOS, upstream RANKL/RANK and downstream nuclear factor of activated T-cells cytoplasmic 1 (NFATC1), and osteoprotegerin OPG. Result:The molecular docking score were -11.86, -11.38, -12.34 kcal·mol-1, and there might be multiple binding sites between TBII as well as RANKL, RANK and C-FOS. Compared with the control group, the content of TRAP in model group increased significantly (P<0.01), and compared with model group, the content of TRAP in each administration group decreased significantly (P<0.01), and TBⅡ decreased the content of TRAP in a dose-dependent manner. Compared with the control group, the expressions of RANKL, RANK, C-FOS and NFATC1 increased (P<0.01), whereas the expression of OPG decreased (P<0.01) in model group. Compared with model group, the expressions of RANKL, RANK, C-FOS and NFATC1 decreased (P<0.01), while the expression of OPG increased (P<0.01) in each administration group. Conclusion:TBⅡ may inhibit the differentiation of osteoclast precursors into osteoclasts, inhibit osteoclast activity, reduce bone resorption and improve osteoporosis by regulating RANKL/RANK/C-FOS signal pathway.

miR-324-5p inhibits lipopolysaccharide-induced proliferation of rat glomerular mesangial cells by regulating the Syk/Ras/c-fos pathway / 南方医科大学学报

OBJECTIVE@#To investigate the effect of miR-324-5p on the proliferation of rat glomerular mesangial (HBZY-1) cells and the role of Syk/Ras/c-fos signaling pathway in mediating this effect.@*METHODS@#HBZY-1 cells cultured in vitro were transiently transfected with miR-324-5p mimics or miR-324-5p-mimics-NC followed by treatment with lipopolysaccharide (LPS). MTT assay was used to detect the proliferation activity of HBZY-1 cells, and RT-qPCR was used to detect the expressions of miR-324-5p and the mRNA expressions of Syk, Ras, MEK1/2, ERK1/2 and c-fos mRNA. The protein expressions of p-Syk, Ras, p-MEK1/2, p-ERK1/2 and c-Fos were detected by Western blotting and immunofluorescence assay.@*RESULTS@#MTT assay showed that exposure to LPS significantly enhanced the proliferative activity of HBZY-1 cells. Compared with the cells treated with LPS and LPS + mimics NC, the cells transfected with miR-324-5p mimics prior to LPS exposure exhibited significantly lowered proliferative activity. Transfection with miR-324-5p mimics significantly lowered the mRNA expressions of Syk, Ras, MEK1/2, ERK1/2 and c-fos and the protein expressions of p-Syk, Ras, MEK1/2, ERK1/2 and c-Fos (@*CONCLUSIONS@#miR-324-5p can inhibit the proliferation of rat chronic glomerulonephritis cells induced by LPS by inhibiting Syk/Ras/c-fos signaling pathway and may potentially serve as a diagnostic indicator and a therapeutic target for chronic glomerulonephritis.

Effect of FIGF overexpression on liver cells transforming to insulin-producing cells

Limitation in the number of insulin-producing pancreatic b-cells is a typical feature of diabetes. It has been indicated thatactivating pancreatic transcription factors can promote the transformation of hepatocytes into insulin-secreting b-like cells,indicating that direct hepatocyte differentiation seems promising as a treatment for diabetes. Nevertheless, the reprogramming efficiency still remains low. Our previous study found that the expression of c-fos-induced growth factor (FIGF)was increased in the pancreatic tissues in partial pancreatectomy mice compared to that in normal mice. Here, we observedthat treatment with Ad-FIGF was found to enhance MafA and Ngn3-induced reprogramming of BNL CL.2 cells to b-likecells with the ability of secreting insulin. And FIGF overexpression increased the levels of histone H3/H4 acetylation atMafA and Ngn3 promoter regions in BNL CL.2 cells. Importantly, in vivo study further confirmed that forced expression ofFIGF facilitated the insulin expression and decreased the blood glucose levels in STZ mice. These results strengthen thepossibility of developing cell-based therapies for diabetes through utilizing b-like cells derived from non-insulin-secretingcells.

Anti-neuropathic pain effect of compound DXL-A-22 / 中国药理学通报

Aim To investigate the anti-neuropathic pain effect of DXL-A-22 and further to explore the potential mechanisms. Methods The anti-neuropathic pain effect was evaluated by chronic constriction injury (CCI) model. The potential anti-neuropathic pain mechanisms of DXL-A-22 was studied by Western blot and qPCR. The acute toxicity was evaluated by ultimate test. Results DXL-A-22 (2,1,0. 5 mg . kg-1 ,i. g.) dose-dependently elevated the mechanical withdrawal threshold (MWT) and the paw withdrawal latency (PWL) in CCI rats (P < 0. 05, P < 0. 01), the percentage of pain threshold elevation (PTE%) and the percentage of Maximal Possible Effect (MPE%) was 108%,86%,71% and 77%,56%,43% respectively on day 7 post-operation. DXL-A-22 (2 mg . kg-1 ,i. g.) significantly reduced the expression of p-CaMK II α, p-CREB, p-JAK2, p-STAT3 proteins and TNF-α mRNA, c-Fos mRNA in DRG (P < 0. 05, P < 0.01), and the percent inhibition was 37%, 48%, 35%,58%, 39% and 32% respectively. The expression of TNF-α mRNA and c-Fos mRNA in spinal pord was reduced by 47% and 72% respectively in CCI rats (P <0. 01). Acute toxicity test showed that DXL-A-22 had no obvious toxicity reaction. Conclusions Spirocyclopiperazinium salt compound DXL-A-22 exerts significant antinociceptive effect on CCI model. The anti-neuropathic pain effect of DXL-A-22 may be related to the inhibition of CaMK II α/CREB and JAK2/STAT3 signaling pathways, and the inhibition of the mRNA expression of TNF-α and c-Fos.

Illuminating the Activated Brain: Emerging Activity-Dependent Tools to Capture and Control Functional Neural Circuits / 神经科学通报·英文版

Immediate-early genes (IEGs) have long been used to visualize neural activations induced by sensory and behavioral stimuli. Recent advances in imaging techniques have made it possible to use endogenous IEG signals to visualize and discriminate neural ensembles activated by multiple stimuli, and to map whole-brain-scale neural activation at single-neuron resolution. In addition, a collection of IEG-dependent molecular tools has been developed that can be used to complement the labeling of endogenous IEG genes and, especially, to manipulate activated neural ensembles in order to reveal the circuits and mechanisms underlying different behaviors. Here, we review these techniques and tools in terms of their utility in studying functional neural circuits. In addition, we provide an experimental strategy to measure the signal-to-noise ratio of IEG-dependent molecular tools, for evaluating their suitability for investigating relevant circuits and behaviors.

Alpha-adrenergic receptors in PVT neurons promote recovery from propofol anesthesia / 医学研究生学报

Objective The locus coeruleus noradrenergic system regulates the recovery process of general anesthesia, but its mechanism remains unclear. The locus coeruleus has a large amount of projection to the paraventricular nucleus of the thalamus (PVT). This study was to investigate the effect of the α-noradrenergic receptor in PVT neurons in propofol anesthesia. Methods The immunofluorescence technique was used for comparison of the c-fos expression in the PVT neurons collected from male SD rats under propofol anesthesia (the PA group, n = 4) or no anesthesia (the non-PA group, n = 4) and observation of the activity of PVT neurons. PVT microinjection models were established in 40 rats and randomized into four groups of equal number: noradrenaline, phentolamine, propranolol, and isotonic saline. Under propofol anesthesia, the animals received microinjection of noradrenaline, phentolamine, propranolol, and isotonic saline at 1 μL into the PVT, respectively, and were observed for the time of recovery of righting reflex (RORR) and the δ (1-4 Hz), θ (4-8 Hz), α (8-12 Hz), β (12-25 Hz) and γ waves (25-60 Hz) on EEG before and after microinjection. Results The expression of c-fos was significantly reduced in the PA group compared with that in the non-PA control. The Ca2+ signals in the PVT were significantly increased during the propofol induction of the loss of righting reflex (LORR), but decreased in the early stage of and during propofol anesthesia (P < 0.05), and remarkably increased at the emergence of and during RORR (P < 0.05). In comparison with the isotonic saline control, the noradrenaline group showed markedly shortened time of RORR (837.8 s vs 647.7 s, P < 0.05), reduced rate of δ waves (P < 0.05) and elevated rate of β waves (P < 0.05), while the phentolamine group exhibited prolonged time of RORR (837.8 s vs 1045.1 s, P < 0.05) and increased rate of δ waves after microinjection (P < 0.05). Conclusion The α-noradrenergic receptors in PVT neurons play a critical role in promoting recovery from propofol anesthesia.

Changes in expression of c-fos protein in spinal cord in a rat model of oxycodone dependence or withdrawal response / 中华麻醉学杂志

Objective To evaluate the changes in the expression of c-fos protein in the spinal cord in a rat model of oxycodone dependence or withdrawal response.Methods Thirty SPF adult male Sprague-Dawley rats,aged 6-8 weeks,weighing 180-220 g,were divided into 3 groups (n=10 each) using a random number table method:normal saline group (group NS),oxycodone dependence group (group OD),and oxycodone withdrawal group (group OW).In OD and OW groups,oxycodone was injected subcutaneously in back,5 days in total,with the dose of 2,3,4,5 and 6 mg/kg in turn,3 times a day (8:00/15:00/22:00).The equal volume of normal saline was given instead in group NS.The mechanical paw withdrawal threshold was measured at 3 days before administration and 30 min after the last administration every day.The oxycodone withdrawal was induced by intraperitoneal injection of naloxone 4 mg/kg at 8 h after the last administration of oxycodone on 5th day in group OW.The withdrawal response scores and range of weight changes were recorded within 15 min after giving naloxone or normal saline in NS and OW groups.Spinal cord tissues were collected at 1 h after the last administration on 5th day in group OD and at 1 h after giving normal saline or naloxone on 5th day in NS and OW groups for determination of the expression of c-fos protein by Western blot.Results Compared with group NS,the mechanical paw withdrawal threshold was significantly increased on 1 and 2 days after administration,and the expression of c-fos protein in the spinal cord was up-regulated in OD and OW groups,and withdrawal response scores were significantly increased,and the range of weight change was increased in group OW (P＜0.05).The expression of c-fos protein was significantly down-regulated in group OW as compared with group OD (P＜0.05).Conclusion Oxycodone dependence or withdrawal response may be related to the expression of c-fos protein in the spinal cord of rats,and the expression is up-regulated during oxycodone dependence,while down-regulated during oxycodone withdrawal.

Synsepalum dulcificum extracts exhibit cytotoxic activity on human colorectal cancer cells and upregulate c-fos and c-jun early apoptotic gene expression

Objective:To explore cytotoxicity of Synsepalum dulcificum (S.dulcificum) Daniell (Sapotaceae) on human colon cancer (HCT-116 and HT-29),human monocytic leukemia (THP-1) and normal (HDFn) cell lines,and its effect on the expression of early apoptotic genes,c-fos and c-jun.Methods:Leaf,stem and berry of S.dulcificum were separately extracted by using 2 solvents,10％ ethanol (EtOH) and 80％ methanol (MeOH).PrestoBlue(R)cell viability assay and qRT-PCR assay were conducted to examine the above objectives respectively.Results:Stem MeOH,stem EtOH,and berry EtOH extracts of S.dulcificum were cytotoxic to HCT-116 and HT-29 human colon cancer cells.For HCT-116,IC50 values of these 3 extracts were not significantly different (P＞0.05) from that of the positive control bleomycin (IC5o of 33.57 μg/mL),while for HT-29,IC5o values of these 3 extracts were significantly lower (P＜0.05) than that of bleomycin (IC50 of 25.24 μg/mL).None of the extracts were cytotoxic to the THP-1 monocytic leukemia cells and HDFn normal human dermal fibroblasts.For both HCT-116 and HT-29,these extracts significantly up-regulated (P＜0.05) the expression of c-fos and c-jun compared to the untreated negative control.Conclusions:The results of this study suggest that cytotoxicity of stem MeOH,stem EtOH,and berry EtOH extracts of S.dulcificum on HCT-116 and HT-29 colon cancer cells is due to the induced apoptosis which is caused by the up-regulation of the expression of early apoptotic genes,c-fos and c-jun.

c-Fos expression in visual cortex of infant rhesus monkeys with myopia induced by hyperopic defocus / 中华实验眼科杂志

Objective To observe c-Fos expression in visual cortex of infant rhesus monkeys with myopia induced by hyperopic defocus and preliminarily investigate the possibility of visual cortex participating in myopia. Methods Eight SPF grade healthy infant rhesus monkeys aged 20 to 30 days were randomly divided into hyperopic defocused group and control group,4 monkeys for each group. The monkeys in hyperopic defocused group wore -3 D spectacle lenses. The monkeys in control group wore 0 D lenses. The monkeys' refractive error,corneal topography, vitreous chamber depth were measured at the start of lens wear and at 2,4,6,8,12 weeks post-treatment. At 12 weeks post-treatment,the visual cortex tissues were removed for c-Fos protein measurement by immunohistochemistry and Western blot assays. The results were analyzed semiquantitatively to compare the differences of c-Fos expression between hyperopic defocused group and control group. The use and care of the animals complied with Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission. This study protocol was approved by Ethic Committee of Zhongshan Ophthalmic Center ( No. 2013-014). Results After 12 weeks'lens wear,the vitreous chamber elongation amplitude of hyperopic defocused group monkeys was more obvious than that of the control group ([0.93±0.24]mm vs. [0.72±0.09]mm;t=2.292,P=0.047). The decrease of hyperopic degrees of hyperopic defocused group monkeys was more obvious than that of the control group ([-3.23± 1.36]D vs. [-1.55±0.52]D;t=-3.273,P=0.006). The eyes of hyperopic defocused group monkeys appeared a remarkable myopic shift after treatment. The number of c-Fos immunoreactive neurons was less in the hyperopic defocused group than that in the control group,with a statistically significant difference between them ([1 843±191]/mm2vs. [2 296±503]/mm2;t=2.381,P=0.041). Western blot assay showed that the optical density of c-Fos protein in the hyperopic defocused group was significantly less than that in the control group (0.50±0.17 vs. 0.99± 0.22;t=-4.982,P<0.01). Conclusions Hyperopic defocus,as an abnormal visual stimulus,can induce the onset of myopia in infant rhesus monkeys and inhibit c-Fos expression in visual cortex. Visual cortex may participate in myopia induced by hyperopic defocus.

Inhibitory effects of solid lipid nanoparticles of rhynchophylline on proliferation of rat vasculars mooth muscle cells induced by TGF-β1 / 国际中医中药杂志

Objective To observe the effect of Rhy-SLN on the proliferation of rat vascular smooth muscle cells (VSMC) induced by TGF-β1, and explore the mechanism. Methods The primary culture of rat thoracic aortic vascular smooth muscle cells was studied by tissue block culture method. The cells were divided into the control group, TGF-β1 group, TGF-β1+ the high, medium and low dosage groups of Rhy-SLN. In addition to the control group, the cells of the other groups were involved in the intervention of TGF-β1 of 20 g/L, and the high, medium and low dosage groups of Rhy-SLN cells were involved in the intervention of 25, 50, 100 mg/L of the hook teng solid lipid nanoparticles. After 24 hours of culture, MTT assay was used to detect cell proliferation inhibition rate in each group, and the cell cycle was detected by flow cytometry. The expression of c-myc and c-Fos protein in each group was detected by Western blot method. Results Compared with the TGF-β1 group, the absorbance value (0.457 ± 0.046 vs. 0.975 ± 0.049) of TGF-β1+ rhy-sln high dose group significantly decreased (P<0.01); the number of S phase cells (15.87% ± 2.47%, 15.23% ± 1.69%, 17.02% ± 2.87% vs.38.58% ± 2.68%)of TGF-β1+rhy-sln in each dose group significantly decreased(P<0.01);The c-myc(48.65 ±3.65,50.69 ± 4.16,55.29 ± 3.67 vs.68.21 ± 3.25)and c-Fos(38.78 ± 4.25,43.56 ± 3.69,46.58 ± 3.57 vs.66.54 ± 4.09) of the TGF-β1+ rhy-sln each dose group significantly decreased (P<0.01). Conclusions The Rhy-SLN can inhibit the proliferation of VSMC in rats induced by TGF-β1.Its mechanism is related to the conversion of G0/G1 phase to the S phase and the expression of the reduction of c-myc and c-fos protein.

Add-on Therapy With the α-Blockers Tamsulosin and Naftopidil Improves Voiding Function by Enhancing Neuronal Activity in Prostatic Hyperplasia Rats / 대한배뇨장애요실금학회지

PURPOSE: Benign prostatic hyperplasia (BPH) impacts quality of life in men by causing lower urinary tract symptoms. α1-Adrenoceptor (α1-AR) blockers improve lower urinary tract symptoms. We investigated the efficacy of add-on therapy with α1-AR blockers on BPH rats. METHODS: Rats in the drug-treated groups were orally administered each drug once a day for 30 days after orchiectomy. To induce BPH, rats were castrated and testosterone (20 mg/kg) was injected subcutaneously once per day for 30 days. Cystometry was conducted to measure voiding contraction pressure and the interval contraction time, immunohistochemistry was performed to measure c-Fos and nerve growth factor (NGF) expression in the neuronal voiding centers, and nicotinamide adenine dinucleotide phosphate-diaphorase histochemistry was used to measure nitric oxide synthase (NOS) expression. RESULTS: Orchiectomy and testosterone injection decreased voiding contraction pressure and the interval contraction time, suggesting BPH symptoms. Voiding contraction pressure and the interval contraction time were greater in the group that received the combination treatment (tamsulosin with naftopidil) than in the tamsulosin monotherapy or naftopidil monotherapy groups. c-Fos, NGF, and NOS expression in the neuronal voiding centers was enhanced by BPH induction. c-Fos, NGF, and NOS expression was suppressed by the combination treatment (tamsulosin with naftopidil) to a greater extent than was the case for tamsulosin monotherapy or naftopidil monotherapy. CONCLUSIONS: Combination therapy of tamsulosin and naftopidil showed greater efficacy for the treatment of BPH than tamsulosin monotherapy or naftopidil monotherapy; therefore, combination therapy can be considered as a novel therapeutic method for BPH.

Effects of dexmedetomidine on expression of c-fos in hippocampus and dentate gyrus in a rat model of endotoxic shock / 中华麻醉学杂志

Objective To evaluate the effects of dexmedetomidine on the expression of c-fos in hippocampus and dentate gyrus in a rat model of endotoxic shock.Methods Thirty-five pathogen-free healthy male Sprague-Dawley rats,aged 3-4 months,weighing 250-300 g,were divided into 5 groups (n =7 each) using a random number table:normal saline group (group NS),dexmedetomidine group (group D),endotoxic shock group (group ES),low-dose dexmedetomidine plus lipopolysaccharide (LPS) group (group LD) and high-dose dexmedetomidine plus LPS group (group HD).Dexmedetomidine 0.5 μg/kg was injected via the tail vein in D and LD groups,and dexmedetomidine 4.5 μg/kg was given in group HD.Normal saline 0.5 ml/kg was injected in NS and ES groups,5 min later normal saline 0.5 ml/kg was injected in NS and D groups and LPS 5 mg/kg was injected in the other groups,and the injection time was 10 min in all groups.Rats were sacrificed at 6 h after LPS injection,brains were removed,and the hippocampus and dentate gyrus were isolated for detection of the expression of c-fos by immunohistochemistry.Results Compared with group NS or group D,the expression of c-fos in the hippocampus and dentate gyrus was significantly up-regulated in group ES (P＜0.05).Compared with group LPS,the expression of c-fos in the hippocampus and dentate gyrus was significantly down-regulated in LD and HD groups (P＜0.05).Compared with group LD,the expression of c-fos in hippocampal CA1 and CA3 areas was significantly down-regulated in group HD (P＜0.05).Conclusion The neuroprotective mechanism of dexmedetomidine is related to inhibiting the up-regulated expression of c-fos in the hippocampus and dentate gyrus in a rat model of endotoxic shock.

Synsepalum dulcificum extracts exhibit cytotoxic activity on human colorectal cancer cells and upregulate c-fos and c-jun early apoptotic gene expression

Objective: To explore cytotoxicity of Synsepalum dulcificum (S. dulcificum) Daniell (Sapotaceae) on human colon cancer (HCT-116 and HT-29), human monocytic leukemia (THP-1) and normal (HDFn) cell lines, and its effect on the expression of early apoptotic genes, c-fos and c-jun. Methods: Leaf, stem and berry of S. dulcificum were separately extracted by using 2 solvents, 10% ethanol (EtOH) and 80% methanol (MeOH). PrestoBlue® cell viability assay and qRT-PCR assay were conducted to examine the above objectives respectively. Results: Stem MeOH, stem EtOH, and berry EtOH extracts of S. dulcificum were cytotoxic to HCT-116 and HT-29 human colon cancer cells. For HCT-116, IC