Engineered interleukin-6-derived cytokines recruit artificial receptor complexes and disclose CNTF signaling via the OSMR.

Ciliary neurotrophic factor (CNTF) activates cells via the non-signaling α-receptor CNTF receptor (CNTFR) and the two signaling ß-receptors glycoprotein 130 (gp130) and leukemia inhibitory factor receptor (LIFR). The CNTF derivate, Axokine, was protective against obesity and insulin resistance, but clinical development was halted by the emergence of CNTF antibodies. The chimeric cytokine IC7 used the framework of interleukin (IL-)6 with the LIFR-binding site from CNTF to activate cells via IL-6R:gp130:LIFR complexes. Similar to CNTF/Axokine, IC7 protected mice from obesity and insulin resistance. Here, we developed CNTF-independent chimeras that specifically target the IL-6R:gp130:LIFR complex. In GIL-6 and GIO-6, we transferred the LIFR binding site from LIF or OSM to IL-6, respectively. While GIO-6 signals via gp130:IL-6R:LIFR and gp130:IL-6R:OSMR complexes, GIL-6 selectively activates the IL-6R:gp130:LIFR receptor complex. By re-evaluation of IC7 and CNTF, we discovered the Oncostatin M receptor (OSMR) as an alternative non-canonical high-affinity receptor leading to IL-6R:OSMR:gp130 and CNTFR:OSMR:gp130 receptor complexes, respectively. The discovery of OSMR as an alternative high-affinity receptor for IC7 and CNTF designates GIL-6 as the first truly selective IL-6R:gp130:LIFR cytokine, whereas GIO-6 is a CNTF-free alternative for IC7.

Ciliary neurotrophic factor promotes the development of homocysteine-induced vascular endothelial injury through inflammation mediated by the JAK2/STAT3 signaling pathway.

Elevated homocysteine (Hcy) levels have been recognized as significant risk factor for cardiovascular and cerebrovascular diseases, closely related to endothelial injury. While expression of Ciliary Neurotrophic Factor (CNTF) significantly increases during Hcy-induced vascular endothelial cell injury, the precise molecular pathways through which CNTF operates remain to be clarified. To induce vascular endothelial cell injury, human umbilical vein endothelial cells (HUVECs) were treated with Hcy. Cell viability and apoptosis in HUVECs were assessed using the CCK-8 assay and flow cytometry. Western blot analysis determined the expression levels of the JAK2-STAT3 pathway, inflammation-related factors (IL-1ß, NLRP3, ICAM-1, VCAM-1), and apoptosis-related factors (cleaved Caspase-3 and Bax). Immunofluorescence staining and western blotting were employed to examine CD31 and α-SMA expression. Knockdown of CNTF was achieved using lentiviral interference, and its effects on inflammation and cell injury were evaluated. Chromatin immunoprecipitation (ChIP) and dual luciferase reporter analysis were conducted to investigate the interaction between the MAFK and CNTF promoters. Our results indicated that Hcy induced high expression of CNTF and activated the JAK2-STAT3 signaling pathway, thereby upregulating factors associated with inflammation and cell apoptosis. Inhibiting CNTF alleviated Hcy-induced inflammation and cell injury. MAFK was identified as a transcription factor promoting CNTF transcription, and its overexpression exacerbated inflammation and cell injury in Hcy-treated HUVECs through the CNTF-JAK2-STAT3 axis, which could be reversed by knocking down CNTF. Activation of MAFK leads to CNTF upregulation, which activates the JAK2-STAT3 signaling pathway, regulating inflammation and inducing injury in Hcy-exposed vascular endothelial cells. Targeting CNTF or its upstream regulator MAFK may represent potential therapeutic strategies for mitigating endothelial dysfunction associated with hyperhomocysteinemia and cardiovascular diseases.

GDE3 regulates oligodendrocyte precursor proliferation via release of soluble CNTFRα.

Oligodendrocyte development is tightly controlled by extrinsic signals; however, mechanisms that modulate cellular responses to these factors remain unclear. Six-transmembrane glycerophosphodiester phosphodiesterases (GDEs) are emerging as central regulators of cellular differentiation via their ability to shed glycosylphosphatidylinositol (GPI)-anchored proteins from the cell surface. We show here that GDE3 controls the pace of oligodendrocyte generation by negatively regulating oligodendrocyte precursor cell (OPC) proliferation. GDE3 inhibits OPC proliferation by stimulating ciliary neurotrophic factor (CNTF)-mediated signaling through release of CNTFRα, the ligand-binding component of the CNTF-receptor multiprotein complex, which can function as a soluble factor to activate CNTF signaling. GDE3 releases soluble CNTFRα by GPI-anchor cleavage from the plasma membrane and from extracellular vesicles (EVs) after co-recruitment of CNTFRα in EVs. These studies uncover new physiological roles for GDE3 in gliogenesis and identify GDE3 as a key regulator of CNTF-dependent regulation of OPC proliferation through release of CNTFRα.

Vanadium inhalation effects on the corneal ciliary neurotrophic factor (CNTF): study in a murine model.

PURPOSE: Air pollution is a public health problem caused by predatory human activities and the indiscriminate burning of fossil fuels that liberate particulate matter (PM) into the atmosphere. Vanadium (V) adheres to them and reaches the bloodstream and different organs such as the eye when inhaled. Another way to reach the eye is by direct contact, and the cornea is the first layer exposed. Ciliary neurotrophic factor (CNTF) is secreted by the corneal nerves and some of its functions include self-renewal maintenance and wound healing by the activation of STAT3. Previous reports from our group indicate the activation of STAT3 after the inhalation of V, adhered to PM. OBJECTIVE: To analyse the effect of V inhalation in the expression of CNTF. Method: CD-1 male mice were exposed for 4 and 8 weeks to V inhalation. The eyes were removed, and the corneas were processed for immunohistochemistry for CNTF and analysed by densitometry. The same slides were used to evaluate histological modifications and to measure the corneas' anterior epithelial and endothelial thickness. RESULTS: A decrease in CNTF expression in the anterior epithelium in the 8th week, as well as an increase in the endothelial and corneal thickness and disarray of all the layers of the anterior epithelium. CONCLUSION: V inhalation disturbs the architecture of the cornea and modifies the presence of CNTF which might modify the renewal of the corneas after exposure to PM air pollution.

[Therapeutic possibilities of stimulating reparative neurogenesis in patients with glaucoma who have recovered from a coronavirus infection]. / Terapevticheskie vozmozhnosti stimulyatsii reparativnogo neirogeneza u patsientov s glaukomoi, perenesshikh koronavirusnuyu infektsiyu.

In patients with glaucoma, the neuroplasticity of retinal cells, their axons and neuroglial elements is pathogenetically reduced, including due to a decrease in the concentration of neurotrophic factors. Coronavirus infections contribute to the damage processes, causing apoptosis of retinal and optic nerve cells. In this regard, the possibility of pharmacological stimulation of the production of these peptides through energy potentiation of the cell mitochondria function, reduction of oxidative stress severity and activation of interneuronal transduction system becomes relevant. PURPOSE: This study aimed to conduct a comprehensive diagnosis of the severity of oxidative stress, identify changes in the neuroplasticity and reparative ability of the retina in patients with primary open-angle glaucoma (POAG) who have recovered after a coronavirus infection, and are undergoing therapy with the complex drug Cytoflavin. MATERIAL AND METHODS: The study included 40 patients (mean age 57.2±3.6 years) with advanced POAG compensated by hypotensive agents; all of them recovered from moderate Covid-19 30 to 90 days prior to inclusion in the study. Twenty patients of the main group received therapy with the complex drug Cytoflavin, 20 other patients comprised the control group. In the comparison groups, the concentration of BDNF and CNTF in blood serum (SC) was determined by enzyme-linked immunosorbent assay (ELISA). Overall assessment of oxidative stress was done by high performance liquid chromatography. Studies of the functional activity of the retina were performed using the Tomey EP 1000 electroretinograph according to the standard method. RESULTS AND DISCUSSION: Retinal photosensitivity significantly improved in patients of the main group taking the complex drug Cytoflavin (mD mean after treatment increased from -7.34±0.62 dB to -4.52±0.12 dB (p>0.001), PSD mean decreased from 6.23±0.21dB to 4.27±0.13 dB (p>0.001)); the neural activity of the retina improved according to PERG (the amplitudes of the P50 and N95 components increased from 0.92±0.04 µv to 1.65±0.01 µv and from 1.83±0.06 µv to 2.68±0.01 µv, respectively (p>0.001), the latency of the P50 and N95 components decreased from 53.40±2.51 ms to 49.37±2.22 ms and from 112.40±5.23 ms to 107.4±8.11ms, respectively (p>0.001); the concentration of BDNF increased (from 18.65±5.32 ng/ml to 20.23±4.05 ng/ml (p>0.001)) and the concentration of CNTF in the blood serum decreased (from 3.99±0.37 pg/ml to 1.85±0.02pg/ml (p>0.001)), the severity of oxidative stress decreased (the indicator of oxidative stress decreased by 1.4 times after treatment p>0.001) and the content of antioxidant protection indicators increased: the indicator of antioxidant protection of blood serum increased by 1.4 times, the concentration of superoxide dismutase - by 1.9 times (p>0.001), glutathione peroxidase - by 1.4 times (p>0.001), coenzyme Q10 - by 4.5 times (p>0.001). CONCLUSION: The obtained data can be used to determine the risk of progression of glaucomatous optic neuropathy in patients with glaucoma who have had a coronavirus infection.

CNTF induces Clcf1 in astrocytes to promote the differentiation of oligodendrocyte precursor cells.

NSCs play an essential role in the regeneration process of the central nervous system. However, due to the influence of the harsh pathological microenvironment, the viability of neural stem cells is limited, and the therapeutic effect needs improvement. Previous studies have found that stem cells overexpressing ciliary neurotrophic factor (CNTF) have apparent therapeutic effects on remyelination, but the specific mechanism of action still needs to be further explored. We found that astrocytes, the most numerous groups in the CNS, exhibited a pathological role in the experimental autoimmune encephalomyelitis model, but after stimulation with CNTF-NSCs, a phenotypic switch occurred and induced the neurotrophic factor cardiotrophin-like cytokine 1 (Clcf1) production. Mechanistically, Clcf1 can significantly promote the differentiation of oligodendrocyte precursor cells (OPCs), and the advanced effect can attenuate by the Clcf1 antibody. Therefore, this study was conducted to investigate the pathway by which CNTF-NSCs exert their therapeutic effects by affecting astrocytes. It is expected to identify a potential therapeutic factor, Clcf1, for the treatment of demyelinating diseases.

Modulation of matrix metalloproteases by ciliary neurotrophic factor in human placental development.

Ciliary neurotrophic factor (CNTF) is a pleiotropic cytokine that signals through a receptor complex containing a specific subunit, CNTF receptor α (CNTFRα). The two molecules are constitutively expressed in key structures for human placental growth and differentiation. The possible role of CNTF in enhancing cell proliferation and/or invasion during placental development and remodelling was investigated using HTR-8/SVneo and BeWo cells, taken respectively as cytotrophoblast and syncytiotrophoblast models. In both cell lines, treatment with human recombinant (hr) CNTF activated JAK2/STAT3 signalling and inhibited the ERK pathway. Interestingly, in HTR-8/SVneo cells, 50 ng hrCNTF induced significant downregulation of matrix metalloprotease (MMP)-1 and significant upregulation of MMP-9. Moreover, pharmacological inhibition of JAK2/STAT3 signalling by AG490 and curcumin resulted in MMP-9 downregulation; it activated the ERK signalling pathway and upregulated MMP-1 expression. Collectively, these data suggest a role for CNTF signalling in extravillous cytotrophoblast invasion through the modulation of specific MMPs.

Ciliary Neurotrophic Factor (CNTF) and Its Receptors Signal Regulate Cementoblasts Apoptosis through a Mechanism of ERK1/2 and Caspases Signaling.

Ciliary neurotrophic factor (CNTF) was identified as a survival factor in various types of peripheral and central neurons, glia and non-neural cells. At present, there is no available data on the expression and localization of CNTF-receptors in cementoblasts as well as on the role of exogenous CNTF on this cell line. The purpose of this study was to determine if cementoblasts express CNTF-receptors and analyze the mechanism of its apoptotic regulation effects on cementoblasts. OCCM-30 cementoblasts were cultivated and stimulated kinetically using CNTF protein (NBP2-35168, Novus Biologicals). Quantified transcriptional (RT-qPCR) and translational (WB) products of CNTFRα, IL-6Rα (CD126), LIFR, p-GP130, GP130, p-ERK1/2, ERK1/2, Caspase-8, -9, -3 and cleaved-caspase-3 were evaluated. Immunofluorescence (IF) staining was applied to visualize the localization of the CNTF-receptors within cells. The apoptosis ratio was measured with an Annexin-V FITC/PI kit. The ERK1/2 antagonist (FR180204, Calbiochem) was added for further investigation by flow cytometry analysis. The CNTF-receptor complex (CNTFRα, LIFR, GP130) was functionally up-regulated in cementoblasts while cultivated with exogenous CNTF. CNTF significantly attenuated cell viability and proliferation for long-term stimulation. Flow cytometry analysis shows that CNTF enhanced the apoptosis after prolonged duration. However, after only a short-term period, CNTF halts the apoptosis of cementoblasts. Further studies revealed that CNTF activated phosphorylated GP130 and the anti-apoptotic molecule ERK1/2 signaling to participate in the regulation of the apoptosis ratio of cementoblasts. In conclusion, CNTF elicited the cellular functions through a notable induction of its receptor complex in cementoblasts. CNTF has an inhibitory effect on the cementoblast homeostasis. These data also elucidate a cellular mechanism for an exogenous CNTF-triggered apoptosis regulation in a mechanism of ERK1/2 and caspase signaling and provides insight into the complex cellular responses induced by CNTF in cementoblasts.

Ciliary Neurotrophic Factor (CNTF) Inhibits In Vitro Cementoblast Mineralization and Induces Autophagy, in Part by STAT3/ERK Commitment.

In animal models, the administration of ciliary neurotrophic factor (CNTF) was demonstrated to reduce bone mass and to participate in bone remodeling. Cementoblasts, a cell type embedded in the cementum, are the main cells to produce and mineralize the extracellular matrix. The effect of CNTF on cementoblasts has not yet been addressed. Thus, the goal of this in vitro study was to investigate possible influences of exogenous CNTF on cementogenesis, as well as autophagy regulation and subsequent mechanisms in cementoblasts. Cementoblasts (OCCM-30) were stimulated with exogenous CNTF. Alizarin Red staining was performed to analyze the functional differentiation (mineralization) of OCCM-30 cells. The release of OPG was quantified by ELISA. The expression of cementogenesis markers (RUNX-2, OCN, BMP-7, BSP, and SPON-2) was evaluated by RT-qPCR. Western blotting (WB) was performed for the protein expression of STAT3, COX-2, SHP-2, cPLAα, cPLAß; ERK1/2, P38, and JNK. The autophagic flux was assessed using WB and RT-qPCR analysis of LC3A/B, Beclin-1, and Atg-5, and the autophagosome was investigated by immunofluorescence staining (IF). The ERK1/2 (FR180204) or STAT3 (sc-202818) antagonist was added, and the cellular response was analyzed using flow cytometry. Exogenous CNTF significantly attenuated mineralized nodule formation, impaired OPG release, and downregulated the mRNA levels of RUNX-2, OCN, BMP-7, and BSP. Moreover, CNTF induced the phosphorylation of STAT3 and activated a transient activation of SHP-2, cPLAß, ERK1/2, P38, and JNK protein. CNTF also induced autophagosome formation and promoted autophagy-associated gene and protein expressions. Additionally, the inhibition of ERK1/2 or STAT3 reversed a CNTF-induced mineralization impairment and had regulatory effects on CNTF-induced autophagosome formation. Our data revealed that CNTF acts as a potent inhibitor of cementogenesis, and it can trigger autophagy, in part by ERK1/2 and STAT3 commitment in the cementoblasts. Thus, it may play an important role in inducing or facilitating inflammatory root resorption during orthodontic tooth movement.

Pleiotropic, Unique and Shared Responses Elicited by IL-6 Family Cytokines in Human Vascular Endothelial Cells.

Vascular endothelial cells express glycoprotein 130 (gp130), which is utilized as a signaling receptor by cytokines in the interleukin-6 (IL-6) family. Several IL-6 family cytokines can be found in the circulatory system during physiological or pathological conditions, and may influence endothelial function and response. This study evaluated and compared the cellular and molecular responses induced by IL-6 family cytokines in human endothelial cells. A proteomic analysis showed that IL-6 family cytokines induce the release of a range of proteins from endothelial cells, such as C-C motif chemokine ligand 23, hepatocyte growth factor, and IL-6. Pathway analysis indicated that gp130-signaling in endothelial cells regulates several functions related to angiogenesis and immune cell recruitment. The present investigation also disclosed differences and similarities between different IL-6 family cytokines in their ability to induce protein release and regulate gene expression and intracellular signaling, in regards to which oncostatin M showed the most pronounced effect. Further, this study showed that soluble gp130 preferentially blocks trans-signaling-induced responses, but does not affect responses induced by classic signaling. In conclusion, IL-6 family cytokines induce both specific and overlapping molecular responses in endothelial cells, and regulate genes and proteins involved in angiogenesis and immune cell recruitment.

Elevated Serum Cortisol Levels in Patients with Focal Epilepsy, Depression, and Comorbid Epilepsy and Depression.

BACKGROUND: The hypothalamic-pituitary-adrenal (HPA) axis, inflammatory processes and neurotrophic factor systems are involved in pathogenesis of both epilepsy and depressive disorders. The study aimed to explore these systems in patients with focal epilepsy (PWE, n = 76), epilepsy and comorbid depression (PWCED n = 48), and major depressive disorder (PWMDD, n = 62) compared with healthy controls (HC, n = 78). METHODS: Parameters of the HPA axis, neurotrophic factors, and TNF-α were measured in blood serum along with the hemogram. RESULTS: Serum cortisol level was augmented in PWE, PWCED, and PWMDD compared with HC and was higher in PWMDD than in PWE. Serum cortisol negatively correlated with Mini-Mental State Examination (MMSE) score in PWE, and positively with depression inventory-II (BDI-II) score in PWMDD. Only PWMDD demonstrated elevated plasma ACTH. Serum TNF-α, lymphocytes, and eosinophils were augmented in PWMDD; monocytes elevated in PWE and PWCED, while neutrophils were reduced in PWE and PWMDD. Serum BDNF was decreased in PWE and PWCED, CNTF was elevated in all groups of patients. In PWE, none of above indices depended on epilepsy etiology. CONCLUSIONS: The results confirm the involvement of HPA axis and inflammatory processes in pathogenesis of epilepsy and depression and provide new insights in mechanisms of epilepsy and depression comorbidity.

p70S6K on astrocytes protects dopamine neurons from 1-methyl-4-phenylpyridinium neurotoxicity.

Our recent finding has demonstrated that astrocytes confer neuroprotection by endogenously producing ciliary neurotrophic factor (CNTF) via transient receptor potential vanilloid 1 (TRPV1) in Parkinson's disease (PD). In this study, the possible molecular target for TRPV1-mediated CNTF production and its neuroprotective effects on dopamine neurons were further investigated. For comparison, glial cell-line derived neurotrophic factor (GDNF) was also examined. The results show that TRPV1-ribosomal protein 70 S6 kinase (p70S6K) signaling on astrocytes produces endogenous CNTF in the SN of MPP+ -lesioned rat. By marked contrast, the expression of GDNF on astrocytes is independent of TRPV1-p70S6K signaling. Administration of a TRPV1 agonist, capsaicin, increases levels of phosphorylated p70S6K (p-p70S6K; activation of p70S6K) on astrocytes, resulting in the survival of dopamine neurons and behavioral recovery through endogenous production of CNTF in the MPP+ -lesioned rat model of PD. Immunohistochemical analysis reveals expression of p-p70S6K on astrocytes in the SN of PD patients, indicating relevance to human PD. The present in vivo data is the first to demonstrate that astrocytic TRPV1-p70S6K signaling plays a pivotal role as endogenous neuroprotective, and it may constitute a novel therapeutic target for treating PD.

The effect of exogenous ciliary neurotrophic factor on cell cycle and neural differentiation markers of in vitro model cells: New insights for future therapeutic approaches.

Retinoblastoma is known as childhood rare malignancy of the retina. Ciliary neurotrophic factor (CNTF) was previously found to reduce degeneration and promote retina survival. This work investigated the effects of CNTF supplementation on in-vitro model cells including retinoblastoma (Y79) and adipose-derived mesenchymal stem cells (AMSCs) viability, proliferation, gene expression and cell cycle. A drop of viability was detected in Y79 treated with CNTF in a dose-dependent manner (P < .05). However, the proliferation of AMSCs was increased at lower concentrations of CNTF (5 ng/mL), but declined in higher doses (50 and 100 ng/mL). The BrdU assay confirmed the MTT assay results. Cell cycle was arrested in both Y79 and AMSCs in the G0/G1 phase by CNTF treatment. A considerable down-regulation of Bcl2, CycD1 and N-Myc genes expression (P < .05) inversely, P15 and P21 genes up-regulation in treated Y79 cells was observed. Besides, stemness genes' transcription was reduced in AMSCs (P < .05), and levels of neuronal-specific markers such as neuron-specific enolase (NSE) and neuronal nuclei (NeuN) were increased (P < .05). The findings of this study suggest a promising potential of CNTF in terms of arresting Y79 retinoblastoma cells, and differentiation-inducing to AMSCs, which could be valuable for managing future innovative treatments targeting retinoblastoma. SIGNIFICANCE OF THE STUDY: We demonstrate that CNTF has the potential to reduce proliferation of Y79 cells and induce the cell cycle arrest of them. Also, down-regulation of oncogenes (such as N-Myc) while up-regulation of tumour suppressor genes (such as P21) was detected by exposure of Y79 cells to CNTF. Furthermore, we observed the cell cycle arrest, reduction of stemness gene and up-regulation of neural differentiation markers in AMSCs treated with CNTF. These results support the probable promising effects of CNTF for controlling retinoblastoma.

Algae Oil Treatment Protects Retinal Ganglion Cells (RGCs) via ERK Signaling Pathway in Experimental Optic Nerve Ischemia.

BACKGROUND: We investigated the therapeutic effects and related mechanisms of algae oil (ALG) to protect retinal ganglion cells (RGCs) in a rat model of anterior ischemic optic neuropathy (rAION). METHODS: Rats were daily gavaged with ALG after rAION induction for seven days. The therapeutic effects of ALG on rAION were evaluated using flash visual evoked potentials (FVEPs), retrograde labeling of RGCs, TUNEL assay of the retina, and ED1 staining of optic nerves (ONs). The levels of inducible nitric oxide synthase (iNOS), IL-1ß, TNF-α, Cl-caspase-3, ciliary neurotrophic factor (CNTF), and p-ERK were analyzed by using western blots. RESULTS: Protection of visual function in FVEPs amplitude was noted, with a better preservation of the P1-N2 amplitude in the ALG-treated group (p = 0.032) than in the rAION group. The density of RGCs was 2.4-fold higher in the ALG-treated group compared to that in the rAION group (p < 0.0001). The number of ED1-positive cells in ONs was significantly reduced 4.1-fold in the ALG-treated group compared to those in the rAION group (p = 0.029). The number of apoptotic RGCs was 3.2-fold lower in number in the ALG-treated group (p = 0.001) than that in the rAION group. The ALG treatment inhibited ERK activation to reduce the levels of iNOS, IL-1ß, TNF-α, and Cl-caspase-3 and to increase the level of CNTF in the rAION model. CONCLUSION: The treatment with ALG after rAION induction inhibits ERK activation to provide both anti-inflammatory and antiapoptotic effects in rAION.

Tetragonia tetragonioides Protected against Memory Dysfunction by Elevating Hippocampal Amyloid-ß Deposition through Potentiating Insulin Signaling and Altering Gut Microbiome Composition.

Alzheimer's disease (AD) is a progressive neurodegenerative disease. Herbal medicine may provide efficacious treatments for its prevention and/or cure. This study investigated whether a 70% ethanol extract of Tetragonia tetragonioides Kuntze (TTK; New Zealand spinach) improved the memory deficit by reducing hippocampal amyloid-ß deposition and modulating the gut microbiota in rats with amyloid-ß(25-35) infused into the hippocampus (AD rats) in an AD animal model. The AD rats had cellulose (AD-CON) or TTK (300 mg/kg bw; AD-TTK) in their high-fat diets for seven weeks. Rats with amyloid-ß(35-25) infused into the hippocampus fed an AD-Con diet did not have memory loss (Normal-Con). AD-TTK protected against amyloid-ß deposition compared to AD-Con, but it was higher than Normal-Con. AD-TTK protected against short-term and special memory loss measured by passive avoidance, Y maze, and water maze, compared to AD-Con. Compared to the Normal-Con, AD-Con attenuated hippocampal pCREB → pAkt → pGSK-3ß, which was prevented in the AD-TTK group. Brain-derived neurotrophic factor (BDNF) and ciliary neurotrophic factor (CNTF) mRNA expression decreased in the AD-CON group, and their expression was prevented in the AD-TTK group. Hippocampal TNF-α and IL-1ß mRNA expressions were higher in the AD-Con group than in the Normal-Con, and AD-TTK groups protected against the increase in their expression. The AD-CON group showed an increase in insulin resistance compared to the Normal-Con group and the AD-TTK group showed improvement. AD-Con separated the gut microbiome community compared to the Normal-Con group and AD-TTK overlapped with the normal-Con. The AD-Con group had more Clostridiales, Erysipelotrichales, and Desulfovibrionales than the AD-TKK and Normal-Con group but fewer Lactobacilales and Bacteroidales. In conclusion, the 70% ethanol extract of TTK enhanced the memory function and potentiated hippocampal insulin signaling, reduced insulin resistance, and improved gut microbiota in amyloid-ß-infused rats.

Tau Gene Deletion does not Influence Axonal Regeneration and Retinal Neuron Survival in the Injured Mouse Visual System.

In the present study, we hypothesized that the microtubule-associated protein Tau may influence retinal neuron survival and axonal regeneration after optic nerve injury. To test this hypothesis, the density of retinal ganglion cells was evaluated by immunostaining retinal flat-mounts for RNA-binding protein with multiple splicing (RBPMS) two weeks after optic nerve micro-crush lesion in Tau-deprived (Tau knock-out (KO)) and wild-type (WT) mice. Axon growth was determined on longitudinal sections of optic nerves after anterograde tracing. Our results showed that the number of surviving retinal ganglion cells and growing axons did not significantly vary between WT and Tau KO animals. Moreover, sustained activation of the neuronal growth program with ciliary neurotrophic factor (CNTF) resulted in a similar increase in surviving neurons and in growing axons in WT and Tau KO mice. Taken together, our data suggest that Tau does not influence axonal regeneration or neuronal survival.

NGF and CNTF expression and regulation mechanism by miRNA in acute paralytic strabismus.

BACKGROUND: Nerve growth factor (NGF) and ciliary neurotrophic factor (CNTF) are well-known neurotrophic factors and widely used in the clinical treatment for its promotion effect on peripheral nerve regeneration. And they were also recommended for the acute paralytic strabismus treatment. However, whether the NGF and CNTF have protective effect for the extraocular muscles of acute paralytic strabismus patients is still poorly understood. PURPOSE: In this study, we want to evaluate the biological function of NGF and CNTF on the extraocular muscle cells and reveale the regulation mechanism behind it. METHODS: Firstly, the relative expression of ngf and cntf was assessed by quantitative real-time RT-PCR. Then, the influence of NGF and CNTF on the extraocular muscle cell proliferation was determined by CCK-8. The inflammatory response in muscle cells after NGF and CNTF treatment was evaluated by ELISA and ROS detection. In addition to this, the up-stream regulation of the ngf and cntf expression was also studied. The TargetScan was used for the predication of potential miRNAs targeting with ngf and cntf 30-UTR, which is soon confirmed by luciferase activity assay. RESULTS: all the results in this research indicated that NGF and CNTF could promote the muscle cell proliferation and inhibit the inflammatory levels, then exert protective effect on the muscle cell function. RESULTS: All the results in this research indicated that NGF and CNTF could promote the muscle cell proliferation and inhibit the inflammatory levels, then exert protective effect on the muscle cell function. CONCLUSION: It was conceivable that let 7-5p was the up-stream regulator of ngf and cntf, and let 7-5p might serve as a potential molecular target for acute paralytic strabismus treatment.

Ciliary neurotrophic factor (CNTF) protects retinal cone and rod photoreceptors by suppressing excessive formation of the visual pigments.

The retinal pigment epithelium (RPE)-dependent visual cycle provides 11-cis-retinal to opsins in the photoreceptor outer segments to generate functional visual pigments that initiate phototransduction in response to light stimuli. Both RPE65 isomerase of the visual cycle and the rhodopsin visual pigment have recently been identified as critical players in mediating light-induced retinal degeneration. These findings suggest that the expression and function of RPE65 and rhodopsin need to be coordinately controlled to sustain normal vision and to protect the retina from photodamage. However, the mechanism controlling the development of the retinal visual system remains poorly understood. Here, we show that deficiency in ciliary neurotrophic factor (CNTF) up-regulates the levels of rod and cone opsins accompanied by an increase in the thickness of the outer nuclear layers and the lengths of cone and rod outer segments in the mouse retina. Moreover, retinoid isomerase activity, expression levels of RPE65 and lecithin:retinol acyltransferase (LRAT), which synthesizes the RPE65 substrate, were also significantly increased in the Cntf-/- RPE. Rod a-wave and cone b-wave amplitudes of electroretinograms were increased in Cntf-/- mice, but rod b-wave amplitudes were unchanged compared with those in WT mice. Up-regulated RPE65 and LRAT levels accelerated both the visual cycle rate and recovery rate of rod light sensitivity in Cntf-/- mice. Of note, rods and cones in Cntf-/- mice exhibited hypersusceptibility to light-induced degeneration. These results indicate that CNTF is a common extracellular factor that prevents excessive production of opsins, the photoreceptor outer segments, and 11-cis-retinal to protect rods and cones from photodamage.

Ciliary Neurotrophic Factor (CNTF) Protects Myocardial Cells from Oxygen Glucose Deprivation (OGD)/Re-Oxygenation via Activation of Akt-Nrf2 Signaling.

BACKGROUND/AIMS: Oxygen glucose deprivation (OGD)/re-oxygenation (OGDR) exposure to myocardial cells mimics ischemia-reperfusion injuries. We studied the potential activity of ciliary neurotrophic factor (CNTF) on OGDR-treated myocardial cells. METHODS: CNTF and CNTFR expression were tested by RT-PCR assay and Western blotting assay. Cell viability and death were tested by MTT assay and LDH release assay, respectively. Akt-Nrf2 signalings were tested by Western blotting assay and qPCR assay. RESULTS: CNTF and its receptor CNTFR were functionally expressed in established H9c2 myocardial cells and primary murine myocardiocytes. Pretreatment of CNTF significantly attenuated OGDR-induced viability reduction and death in myocardial cells. Further studies show that in the myocardial cells CNTF activated NF-E2-related factor 2 (Nrf2) signaling to inhibit OGDR-induced reactive oxygen species (ROS) production and programmed necrosis, preventing adenine nucleotide translocator 1 (ANT-1)-p53-cyclophilin D (Cyp-D) mitochondrial association and mitochondrial depolarization. Nrf2 silencing or knockout almost abolished CNTF-induced H9c2 cytoprotection against OGDR. CNTF activated Akt in H9c2 cells and primary murine myocardiocytes. Conversely, Akt blockage by the pharmacological inhibitors not only blocked CNTF-induced Nrf2 Ser-40 phosphorylation and activation, but also nullified anti-OGDR actions by CNTF in myocardial cells. CONCLUSION: CNTF activates Akt-Nrf2 signaling to protect myocardial cells from OGDR.

mTORC1 pathway disruption abrogates the effects of the ciliary neurotrophic factor on energy balance and hypothalamic neuroinflammation.

Ciliary neurotrophic factor (CNTF) potently decreases food intake and body weight in diet-induced obese mice by acting through neuronal circuits and pathways located in the arcuate nucleus (ARC) of the hypothalamus. CNTF also exerts pro-inflammatory actions within the brain. Here we tested whether CNTF modifies energy balance by inducing inflammatory responses in the ARC and whether these effects depend upon the mechanistic target of rapamycin complex 1 (mTORC1) pathway, which regulates both energy metabolism and inflammation. To this purpose, chow- and high fat diet (HFD)- fed mice lacking the S6 kinase 1 (S6K1-/-), a downstream target of mTORC1, and their wild-type (WT) littermates received 12 days continuous intracerebroventricular (icv) infusion of the CNTF analogue axokine (CNTFAx15). Behavioral, metabolic and molecular effects were evaluated. Central chronic administration of CNTFAx15 decreased body weight and feed efficiency in WT mice only, when fed HFD, but not chow. These metabolic effects correlated with increased number of iba-1 positive microglia specifically in the ARC and were accompanied by significant increases of IL-1ß and TNF-α mRNA expression in the hypothalamus. Hypothalamic iNOS and SOCS3 mRNA, molecular markers of pro-inflammatory response, were also increased by CNTFAx15. All these changes were absent in S6K1-/- mice. This study reveals that CNTFAx15 requires a functional S6K1 to modulate energy balance and hypothalamic inflammation in a diet-dependent fashion. Further investigations should determine whether S6K1 is a suitable target for the treatment of pathologies characterized by a high neuroinflammatory state.