Nerve Growth Factor Secretion Research Articles

Regulatory effect of nerve growth factor in α9β1 integrin–dependent progression of glioblastoma

In the present study we described the role of alpha9beta1 integrin in glioblastoma progression following its interaction with nerve growth factor (NGF). The level of expression of alpha9beta1 on astrocytomas is correlated with increased grade of this brain tumor and is highest on glioblastoma, whereas normal astrocytes do not express this integrin. Two glioblastoma cell lines, LN229 and LN18, that are alpha9beta1 integrin positive and negative, respectively, were used for alpha9beta1 integrin-dependent NGF-induced tumor progression. NGF was a significant promoter of promigratory and pro-proliferative activities of glioblastoma cells through direct interaction with alpha9beta1 integrin and activation of MAPK Erk1/2 pathway. The level of NGF increases approximately threefold in the most malignant glioma tissue when compared with normal brain. This increase is related to secretion of NGF by tumor cells. Specific inhibitors of alpha9beta1 integrin or gene silencing inhibited NGF-induced proliferation of LN229 cell line to the level shown by LN18 cells. VLO5 promoted alpha9beta1-dependent programmed cell death by induction of intrinsic apoptosis pathway in cancer cells. LN229 cells were rescued from proapoptotic effect of VLO5 by the presence of NGF. This disintegrin significantly inhibited tumor growth induced by implantation of LN229 cells to the chorioallantoic membrane (CAM) of quail embryonic model, and this inhibitory effect was significantly abolished by the presence of NGF. alpha9beta1 integrin appears to be an interesting target for blocking the progression of malignant gliomas, especially in light of the stimulatory effect of NGF on the development of these tumors and its ability to transfer proapoptotic signals in cancer cells.

Basal forebrain stimulation induces NGF secretion in ipsilateral parietal cortex via nicotinic receptor activation in adult, but not aged rats

The role of cholinergic basal forebrain inputs to cerebral cortex in regulating regional nerve growth factor (NGF) secretion was examined in adult (4–6 months) and aged (29–31 months) rats. Halothane-anesthetized rats received unilateral electrical stimulation of the nucleus basalis of Meynert (NBM). NGF levels were measured by ELISA in samples from a microdialysis probe in the parietal cortex, while measuring blood flow. In adult and aged rats, NBM stimulation (for 100 min) increased blood flow ipsilaterally during stimulation by 55% and 25%, respectively. In adult, but not aged rats, NGF levels were significantly increased ipsilaterally (up to 68%) over prestimulus levels at 200–500 min after stimulation ended. The cellular localization of NGF-like immunoreactivity showed no differences between the cortices with and without NBM stimulation. The NGF response was abolished by the nicotinic blocker, mecamylamine (20 mg/kg iv), but unaffected by the muscarinic blocker, atropine (5 or 25 mg/kg iv). Both drugs reduced the blood flow responses. We conclude that cholinergic inputs to neocortex mediate NGF secretion by cortical neurons via nicotinic receptors. Further, the absence of this response in aged rats suggests a decline in the number or activity of cortical nicotinic receptors.

NGF Gene Expression and Secretion by Canine Adipocytes in Primary Culture: Upregulation by the Inflammatory Mediators LPS and TNFα

Obesity is the commonest nutritional disorder of companion animals. In rodents and humans, white adipose tissue is a major endocrine and secretory organ, releasing adipokines linked to inflammation. In this study, we examined whether nerve growth factor (NGF), a target-derived neurotrophin central to the development/maintenance of sympathetic innervation and an inflammatory response protein, is synthesized and secreted by canine adipocytes. NGF mRNA was detected in each of the major fat depots (the subcutaneous, inguinal, gonadal, perirenal, and falciform ligaments) of dogs at similar levels. Canine adipocytes, differentiated from preadipocytes (inguinal depot) in primary culture, expressed the NGF gene and secreted NGF both pre- and post-differentiation. Treatment of the differentiated adipocytes with LPS resulted in a dramatic increase in NGF mRNA levels (20-fold at 24 h) and in NGF protein in the medium (60-fold at 24 h). The proinflammatory cytokine TNFalpha also led to a substantial increase in NGF mRNA levels (11-fold) and protein secretion (16-fold), while IL-6 had little effect. In contrast, dexamethasone decreased both NGF mRNA levels (80%) and protein release (60%). The PPARgamma agonist rosiglitazone also reduced NGF secretion. These results demonstrate that canine white adipocytes synthesize and secrete NGF, the powerful upregulation by LPS and TNFalpha indicating that the neurotrophin is strongly linked to the inflammatory response in canine WAT. Canine adipocytes appear highly sensitive to inflammatory stimuli.

ABM Clinical Protocol #4: Mastitis

Breastfeeding MedicineVol. 3, No. 3 ABM PROTOCOLSABM Clinical Protocol #4: MastitisThe Academy of Breastfeeding Medicine Protocol CommitteeThe Academy of Breastfeeding Medicine Protocol CommitteeSearch for more papers by this authorPublished Online:8 Sep 2008https://doi.org/10.1089/bfm.2008.9993AboutSectionsPDF/EPUB ToolsPermissionsDownload CitationsTrack CitationsAdd to favorites Back To Publication ShareShare onFacebookTwitterLinked InRedditEmail FiguresReferencesRelatedDetailsCited byRelationship between Acute Mastitis and Constitution of Traditional Chinese Medicine in Chinese Breastfeeding MothersEvidence-Based Complementary and Alternative Medicine, Vol. 2021Influence of Previous COVID-19 and Mastitis Infections on the Secretion of Brain-Derived Neurotrophic Factor and Nerve Growth Factor in Human Milk8 April 2021 | International Journal of Molecular Sciences, Vol. 22, No. 8Lactational Breast Abscesses Caused by Methicillin-Resistant or Methicillin-Sensitive Staphylococcus aureus Infection and Therapeutic Effect of Ultrasound-Guided Aspiration Song-tao Ding, Xiang-ping He, Xiang-jun Ma, Yi Zhang, Xing-xing Liu, and Jing Qin10 July 2020 | Breastfeeding Medicine, Vol. 15, No. 7Negative-Pressure Suction Cannula for Treatment of Lactational Residual Milk Cavity: A Case Series Study Yajun Gao, Xiangjun Ma, Xiangping He, Haifeng Gao, Songtao Ding, Yan Li, and Yi Zhang9 June 2020 | Breastfeeding Medicine, Vol. 15, No. 6Comprehensive evaluation of the risk of lactational mastitis in Chinese women: combined logistic regression analysis with receiver operating characteristic curve20 March 2020 | Bioscience Reports, Vol. 40, No. 3Lactostasis and prevention of lactation mastitis: role of neonatologist and pediatrician9 March 2020 | Meditsinskiy sovet = Medical Council, No. 1ProbioticsBenign Breast Disease in Women19 August 2019 | Deutsches Ärzteblatt internationalDevelopment and Evaluation of a Lactation Rotation for a Pediatric Residency Program20 March 2017 | Journal of Human Lactation, Vol. 33, No. 4Puerperal Complications11 June 2017Management of mastitis and breast engorgement in breastfeeding women30 November 2015 | The Journal of Maternal-Fetal & Neonatal Medicine, Vol. 29, No. 19Negative Suction Drain Through a Mini Periareolar Incision for the Treatment of Lactational Breast Abscess Shortens Hospital Stay and Increases Breastfeeding Rates Jinli Wei, Jiaxin Zhang, and Deyuan Fu15 June 2016 | Breastfeeding Medicine, Vol. 11, No. 5Principles of Breastfeeding and Navigating Its ChallengesPostgraduate Obstetrics & Gynecology, Vol. 34, No. 23Taste of Milk from Inflamed Breasts of Breastfeeding Mothers with Mastitis Evaluated Using a Taste Sensor Michiko Yoshida, Hitomi Shinohara, Toshihiro Sugiyama, Masanori Kumagai, Hajime Muto, and Hideya Kodama24 February 2014 | Breastfeeding Medicine, Vol. 9, No. 2Enabling Women to Achieve Their Breastfeeding GoalsObstetrics & Gynecology, Vol. 123, No. 3Diagnosis and Treatment of Mastitis in Breastfeeding Women6 January 2014 | Journal of Human Lactation, Vol. 30, No. 1A Case Control Study of Bacterial Species and Colony Count in Milk of Breastfeeding Women with Chronic Pain Ann Witt, Mary Jane Mason, Kelly Burgess, Susan Flocke, and Steven Zyzanski23 January 2014 | Breastfeeding Medicine, Vol. 9, No. 1Mastitis in Chinese Breastfeeding Mothers: A Prospective Cohort Study Li Tang, Andy H. Lee, Liqian Qiu, and Colin W. Binns23 January 2014 | Breastfeeding Medicine, Vol. 9, No. 1Postpartum Breast Abscess Caused by Community-Acquired Methicillin-Resistant Staphylococcus aureus in Japan Hideharu Hagiya, Sumiko Shiota, Wakana Sugiyama, and Fumio Otsuka23 January 2014 | Breastfeeding Medicine, Vol. 9, No. 1To Feed or Not to Feed? Case Presentation and Best Practice Guidance for Human Milk Feeding and Group B Streptococcus in Developed Countries18 March 2013 | Journal of Human Lactation, Vol. 29, No. 4Treatments for breast abscesses in breastfeeding women30 April 2013Maternal and infant infections stimulate a rapid leukocyte response in breastmilkClinical & Translational Immunology, Vol. 2, No. 4Dynamics of Breast Milk HIV-1 RNA With Unilateral Mastitis or AbscessJAIDS Journal of Acquired Immune Deficiency Syndromes, Vol. 62, No. 3Circumstances when Breastfeeding is ContraindicatedPediatric Clinics of North America, Vol. 60, No. 1Clinical Protocols for Management of BreastfeedingPediatric Clinics of North America, Vol. 60, No. 1MExtremely Large Breast Abscess in a Breastfeeding Mother17 May 2012 | Journal of Human Lactation, Vol. 28, No. 4Postpartum care aa Includes breastfeeding, contraception, postpartum endometritis, and postpartum wound infection.14 March 2012Breastfeeding: A Clinical Imperative Michelle G. Brenner and E. Stephen Buescher14 December 2011 | Journal of Women's Health, Vol. 20, No. 12Defining Lactation Acuity to Improve Patient Safety and Outcomes28 April 2011 | Journal of Human Lactation, Vol. 27, No. 2Medical Complications of MothersBreastfeeding Health SupervisionPainful Breast Lumps in Nursing MothersAJN, American Journal of Nursing, Vol. 110, No. 12 Volume 3Issue 3Sep 2008 Information© 2008 Mary Ann Liebert, Inc.To cite this article:The Academy of Breastfeeding Medicine Protocol Committee.ABM Clinical Protocol #4: Mastitis.Breastfeeding Medicine.Sep 2008.177-180.http://doi.org/10.1089/bfm.2008.9993Published in Volume: 3 Issue 3: September 8, 2008PDF download

Panaxydol treatment enhances the biological properties of Schwann cells in vitro

Schwann cells (SCs), the glial cells of the peripheral nerve system, play a key role in the regeneration of injured peripheral nerves. However, problems with the use of SCs to repair peripheral nerves include attenuated biologic properties and impaired function with ageing. Panaxydol (PND) effectively protects neurons against injury in degenerative diseases. We investigated the protective role of PND in SCs through immunocytochemistry and ELISA assay. PND promoted the expression and secretion of nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF) by SCs in a dose-dependent manner at doses of 2.5–20 and 5.0–20 μM, respectively. The effects on both factors were maximal at 10 μM. PND also enhanced the synthesis of actin, a key component of the cytoskeleton. When we examined mitochondria in SCs with probes marked with rhodamine-123, fluorescence intensity was stronger in the PND group than in a control group, indicating a stabilized mitochondrial transmembrane potential. PND modified cytoskeleton dynamics and induced SCs to secrete and express neurotrophic factors (NTFs), and to resist high energy consumption in a dose-dependent manner. It exerted its maximum effect at 10 μM. PND treatment of SCs might be promising strategies for the application of these cells in repairing PNS injury by enhancing the biological properties.

Regulatory effect of nerve growth factor in 9 1 integrin-dependent progression of glioblastoma

In the present study we described the role of alpha9beta1 integrin in glioblastoma progression following its interaction with nerve growth factor (NGF). The level of expression of alpha9beta1 on astrocytomas is correlated with increased grade of this brain tumor and is highest on glioblastoma, whereas normal astrocytes do not express this integrin. Two glioblastoma cell lines, LN229 and LN18, that are alpha9beta1 integrin positive and negative, respectively, were used for alpha9beta1 integrin-dependent NGF-induced tumor progression. NGF was a significant promoter of promigratory and pro-proliferative activities of glioblastoma cells through direct interaction with alpha9beta1 integrin and activation of MAPK Erk1/2 pathway. The level of NGF increases approximately threefold in the most malignant glioma tissue when compared with normal brain. This increase is related to secretion of NGF by tumor cells. Specific inhibitors of alpha9beta1 integrin or gene silencing inhibited NGF-induced proliferation of LN229 cell line to the level shown by LN18 cells. VLO5 promoted alpha9beta1-dependent programmed cell death by induction of intrinsic apoptosis pathway in cancer cells. LN229 cells were rescued from proapoptotic effect of VLO5 by the presence of NGF. This disintegrin significantly inhibited tumor growth induced by implantation of LN229 cells to the chorioallantoic membrane (CAM) of quail embryonic model, and this inhibitory effect was significantly abolished by the presence of NGF. alpha9beta1 integrin appears to be an interesting target for blocking the progression of malignant gliomas, especially in light of the stimulatory effect of NGF on the development of these tumors and its ability to transfer proapoptotic signals in cancer cells.

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Interstitial cystitis (IC) is a disease of mystery. This disease causes increased bladder sensitivity, decreased bladder capacity, and a painful bladder, whether empty or full. Recently, most physicians have concluded that IC is an inflammatory disease that can progress if not treated. However, the effects of treatment aiming at protection of the urothelium have usually been limited. 1 Nickel J.C. Barkin J. Forrest J. et al. Randomized, double-blind, dose-ranging study of pentosan polysulfate sodium for interstitial cystitis. Urology. 2005; 65: 654-658 Abstract Full Text Full Text PDF PubMed Scopus (141) Google Scholar Botulinum toxin type A (BTX-A) injection provides a novel treatment of IC, decreasing bladder sensitivity by reducing the sensory receptors, such as P2X3 and transient receptor potential vanilloid 1, 2 Apostolidis A. Papat R. Yiangou Y. et al. Decreased sensory receptors P2X3 and TRPV1 in suburothelial nerve fibers following intradetrusor injection of botulinum A toxin for human detrusor overactivity. J Urol. 2005; 173: 977-983 Abstract Full Text Full Text PDF Scopus (359) Google Scholar increasing bladder capacity by paralyzing the detrusor, 3 Kuo H.C. Urodynamic evidence of effectiveness of botulinum A toxin injection in treatment of detrusor overactivity refractory to anticholinergic agents. Urology. 2004; 63: 868-872 Abstract Full Text Full Text PDF PubMed Scopus (173) Google Scholar and decreasing bladder pain by suppressing the release of calcitonin gene-related peptide and other neuropeptides. 4 Rapp D.E. Turk K.W. Bales G.T. et al. Botulinum toxin type A inhibits calcitonin gene-related peptide release from isolated rat bladder. J Urol. 2006; 175: 1138-1142 Abstract Full Text Full Text PDF PubMed Scopus (143) Google Scholar The actual mechanism of effective BTX-A treatment has not been elucidated. It is possibly through inhibition of the inflammatory process. This letter-to-editor points out the underlying molecular mechanism of BTX-A affects on the proinflammatory pathway, which further modulates the secretion of nerve growth factor from the bladder wall. By inhibiting the inflammatory process in the suburothelium or urothelium, BTX-A not only might decrease the apoptosis of urothelial cells, but also could reestablish the integrity of the injured urothelium. However, because the inflammatory process in IC might involve the corresponding spinal cords, 5 Cui M. Aoki K.R. Botulinum toxin type A (BTX-A) reduces inflammatory pain in the rat formalin model. Cephalalgia. 2000; 20: 414-418 Crossref Google Scholar the therapeutic result of BTX-A on IC through detrusor injection cannot last for long. Repeated BTX-A injections might provide long-term desensitization to the inflammation of the central nervous system in IC and a better treatment outcome. Intravesical Botulinum Toxin A Injections Plus Hydrodistension Can Reduce Nerve Growth Factor Production and Control Bladder Pain in Interstitial Cystitis: A Molecular MechanismUrologyVol. 72Issue 2PreviewI read with great interest the article by Liu and colleagues.1 This work shows that botulinum toxin markedly reduces nerve growth factor (NGF) compared with a placebo. I would like to complete the discussion of Liu and coworkers1 by suggesting a major route through which botulinum toxin could reduce NGF. Full-Text PDF

Helminth infections associated with multiple sclerosis induce regulatory B cells

To assess the importance of B-cell control during parasite infections in multiple sclerosis (MS) patients. Peripheral blood CD19+ B cells from 12 helminth-infected MS patients, 12 MS patients without infection, 10 patients infected with Trypanosoma cruzi, 8 subjects infected with Paracoccidioides brasiliensis, and 12 healthy control subjects were purified using magnetic cell sorting. Interleukin (IL)-4, IL-6, IL-10, tumor necrosis factor-alpha, lymphotoxin, transforming growth factor-beta, brain-derived neurotrophic factor, and nerve growth factor secretion were evaluated after stimulation with CDw32 L cells and CD40 antibody using enzyme-linked immunosorbent assays. The production of anti-myelin oligodendrocyte glycoprotein IgG and IgM antibodies was evaluated by enzyme-linked immunosorbent spot assays. Cell phenotype was assessed by flow cytometry. Helminth infections in MS patients created a B-cell population producing high levels of IL-10, dampening harmful immune responses through a mechanism mediated, at least in part, by the ICOS-B7RP-1 pathway. The IL-10-producing B-cell phenotype detected expressed high levels of CD1d and was similar to the one observed in mature naive B2 cells (namely, CD11b(-), CD5(-), CD27(-), and IgD+). Moreover, B cells isolated from helminth-infected MS patients also produced greater amounts of brain-derived neurotrophic factor and nerve growth factor compared with those of normal subjects, T. cruzi-infected subjects, P. brasiliensis-infected subjects, or uninfected MS patients, raising the possibility that these cells may exert a neuroprotective effect on the central nervous system. Increased production of B-cell-derived IL-10 and of neurotrophic factors are part of the parasite's regulation of host immunity and can alter the course of MS, potentially explaining environmental-related MS suppression observed in areas with low disease prevalence.

Reduced NGF secretion by Schwann cells under the high glucose condition decreases neurite outgrowth of DRG neurons

BackgroundSchwann cells (SCs) have been supposed to play prominent roles in axonal regeneration under various diseases. Here, to evaluate the direct interaction between SCs and dorsal root ganglion (DRG) neurons under a diabetic condition, the effects of Schwann cell-conditioned media on neurite outgrowth of DRG neurons were investigated. MethodsImmortalized mouse Schwann cells (IMS) were cultured under 5.5 mM glucose (NG) or 30 mM glucose (HG) conditions for 4 days. IMS-conditioned media (IMS-media) were added to the culture media of neurons isolated from 8-week-old DDY mice. Neurons were cultured for 48 h with or without mouse recombinant NGF (mrNGF) or nerve growth factor (NGF) neutralizing antibody. The concentrations of NGF in IMS-media by ELISA and neurite outgrowth by a computed image analysis system were evaluated. ResultsNeurite outgrowth was significantly enhanced by IMS-media (IMS-media (–): 177±177 µm, IMS-media (+): 1648±726). The neurite outgrowth cultured with IMS-media obtained under the HG condition was significantly reduced compared with that under the NG condition (NG: 1474±652, HG: 734±331). The NGF concentrations were significantly lower in IMS-media under the HG condition than in those under the NG condition. The accelerated neurite outgrowth by IMS-media was inhibited by NGF neutralizing antibody. ConclusionsThese results suggest that SCs play important roles in neurite outgrowth of DRG neurons, and that the decreased NGF secretion by SCs under the diabetic condition would cause a defect of axonal regeneration, resulting in the development of diabetic neuropathy.

Enhanced production and secretion of glial cell line-derived neurotrophic factor and nerve growth factor from the skin in atopic dermatitis mouse model

Role of neurotrophic factors including nerve growth factor (NGF) in the mechanism of overgrowth and hypersensitivity of sensory nerve in atopic dermatitis (AD) has been proposed. Glial cell line-derived neurotrophic factor (GDNF) is a member of neurotrophic factors of the nervous systems; however, the role of GDNF in dermatitis is unknown. IL-18 promotes Th2 type allergic condition in skin and various organs in the absence of IL-12. In this report, we evaluated the expression of GDNF in AD and its association with NGF and IL-18. Mice expressing skin-specific IL-18 (KIL18Tg) or caspase-1, an IL-18 converting enzyme, (KCASP1Tg) were used as AD models; GDNF expression was examined by RT-PCR, enzyme immunoassay, and immunohistochemistry. The mRNA expressions of GDNF and NGF were detected in the epidermis and they were increased in the skin of KIL18Tg and KCASP1Tg mice. GDNF protein production in the skin was also elevated in both transgenic mice and mostly expressed at the basal layer of the epidermis as assessed by immunohistochemistry. Furthermore, the number of nerve fibers was increased in KCASP1Tg, suggesting increased cutaneous innervation. The present results suggest that in addition to NGF, elevated production and secretion of GDNF in the skin associated with overproduction of IL-18 may also be a potent causative factor of itching in AD.

Exploring Human Testicular Peritubular Cells: Identification of Secretory Products and Regulation by Tumor Necrosis Factor-α

Testicular peritubular cells are myofibroblastic cells, which represent the major cellular components of the wall of the seminiferous tubules. In men their phenotypic characteristics, including possible secretory activity and regulation, are not well known, in neither normal nor pathologically altered testes. Especially in testes of men with impaired spermatogenesis, the cytoarchitecture of the tubular wall is frequently remodeled and presents fibrotic thickening, increased innervation, and infiltration by macrophages and mast cells. The latter are two sources of TNF-alpha. The purpose of our study was to explore human testicular peritubular cells and mechanisms of their regulation. To this end we primarily studied cultured human testicular peritubular cells (HTPCs), isolated from adult human testes. Having established that HTPCs express TNF-alpha receptors 1 and 2 and respond to recombinant human TNF-alpha by a rapid phosphorylation of ERK1/2, we used complementary approaches, including gene array/RT-PCR studies, Western blotting/immunocytochemistry, and ELISA techniques to study phenotypic characteristics of HTPCs and actions of TNFalpha. We found that HTPCs express the nerve growth factor gene and TNF-alpha-stimulated mRNA levels and secretion of nerve growth factor in a dose- and time-dependent manner. Similarly, monocyte chemoattractant protein-1 was identified as a product of HTPCs, which was regulated by TNF-alpha in a concentration- and time-dependent manner. TNF-alpha furthermore strongly enhanced expression and/or synthesis of other inflammatory molecules, namely IL-6 and cyclooxygenase-2. Active cyclooxygenase-2 is indicated by increased prostaglandin D2 levels. In addition, intercellular adhesion molecule-1, which was not detected at protein level in the absence of TNF-alpha, was induced upon TNF-alpha stimulation. In conclusion, these results provide novel insights into the nature of human peritubular cells, which are able to secrete potent signaling molecules and are regulated by TNF-alpha. These results also hint to an as-yet-unknown role of peritubular cells in normal human testis and involvement in the pathomechanisms associated with impaired spermatogenesis in men.

Submandibular glands in the metabolic syndrome

In addition to their stimulatory action on neuronal differentiation and survival, a variety of neurotrophic factors, such as nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and ciliary neurotrophic factor, exert metabotrophic effects, including improvement of glucose, lipid and energy homeostasis. It was recently reported that plasma levels of both NGF and BDNF are reduced in patients with advanced metabolic syndrome and with acute coronary syndromes, and that NGF tissue content is decreased in human atherosclerotic coronary arteries. Since NGF and BDNF are synthesized, stored, and released by submandibular salivary glands, we investigated the structure and function of these glands. Here we present our scintigraphic and echographic results of submandibular glands of patients with advanced stage of metabolic syndrome: (i) scintigraphic analysis using the radiotracer (99m)Tc-pertechnetate showed an inhibition of salivary gland excretory activity, and (ii) echographic evaluation revealed a parenchymal destruction and a prominent fibrosis of the glands. Both suggestive for the involvement of submandibular glands in decreased secretion of NGF and BDNF as implicated in the pathogenesis of metabolic syndrome. Biomedical Reviews 2007; 18: 65-67.

Nerve Growth Factor Promotes Development of Glucose Induced Insulin Secretion in Rat Neonate Pancreatic Beta Cells by Modulating Calcium Channels

Neonatal javascript:app('lower case beta')ß cells are functionally immature as they secrete less insulin than adults and lack of glucose response. The mechanisms that participate in the functional maturation of these cells are not known. Adult rat javascript:app('lower case beta')ß cells synthesize and secrete nerve growth factor (NGF) and express NGF receptors. NGF increases glucose-induced insulin secretion by modulating electrical activity in adult javascript:app('lower case beta')ß cells. In this work, we explored if NGF is involved in the maturation of glucose-induced insulin secretion coupling in rat neonate javascript:app('lower case beta')ß-cells.Pancreatic javascript:app('lower case beta')ß-cells were cultured for 48 h in control conditions, either with NGF or a neutralizing NGF antibody. We measured glucose-stimulated insulin secretion and whole-cell Ca2+ currents. We also analyzed both synthesis and molecular expression of Ca2+ channel subunits.Compared to controls, NGF-treated neonate javascript:app('lower case beta')ß-cells increased insulin biosynthesis and secretion in response to glucose and increased Ca2+ current density by translocation of LD-type Ca2+ channels to the plasma membrane. This type of secretion is similar to that observed in adults.NGF contributes to in vitro maturation by inducing beta cells to secrete insulin accordant with external glucose concentration, in other words promoting glucose sensitivity. Similar NGF-autocrine mechanisms could be active in vivo, because these effects were blocked by a neutralizing NGF-antibody. Understanding mechanisms that contribute to normal development of javascript:app('lower case beta')ß cell could provide insights to recognize alterations concomitant with type-2 diabetes.

5,19-cyclo-9β,10ξ-androstane-3,17-dione promotes neurotrophic factor biosynthesis in 1321N1 human astrocytoma cells and improves passive avoidance learning impairment

Since neurotrophic factors are essential for neurons to form neuronal networks and maintain neuronal functions, neurotrophic factor-like substances or inducers of neurotrophic factors can be useful for the treatment of serious neuronal diseases such as Alzheimer's and Parkinson's diseases. In the present study, we examined an effect of 5,19-cyclo-9β,10ξ-androstane-3,17-dione (CAD) on neurotrophic factor synthesis in glial cells and scopolamine-induced impairment of learning in mice. 1321N1 human astrocytoma cells promoted secretion of certain neurotrophic factors in response to CAD with no cytotoxicity, which caused dramatic neurite outgrowth in rat pheochromocytoma (PC12) cells. In fact, CAD significantly enhanced nerve growth factor (NGF) secretion and its gene expression in 1321N1 cells, in a time and concentration-dependent manner. Because second messengers such as cAMP, inositol 1,4,5-trisphosphates and Ca 2+ induce NGF gene expression, we measured activities of adenylyl cyclase and phospholipase C and intracellular Ca 2+ concentration in 1321N1 cells. However, CAD changed neither second messenger levels. CAD enhanced the gene expression of proto-oncogene, c-fos that is one of the components of transcription factor (AP-1). In addition to those above, the in vivo effects of CAD were also examined. Although injection of muscarinic receptor antagonist scopolamine impaired passive avoidance learning in mice, pretreatment with CAD significantly reversed the adverse effect in a dose-dependent manner. Taking these results together, CAD has enormous therapeutic potential for serious neuronal diseases.

Cross‐talk between the epidermal growth factor‐like repeats/fibronectin 6–8 repeats domains of Tenascin‐R and microglia modulates neural stem/progenitor cell proliferation and differentiation

Mounting evidence has demonstrated that the microenvironment of stem/progenitor cells plays an important role in their proliferation and commitment to their fate. However, it remains unclear how all elements, such as astrocytes, microglia, extracellular matrix molecules, soluble factors, and their cross-talk interactions in the microenvironments, affect neural stem/progenitor cell fate. This work explored the influences of cross-talk between Tenascin-R (TN-R) and microglia on neural stem/progenitor cell proliferation and differentiation. Our results show that microglia triggered by TN-R distinct domains EGF-like repeats (EGFL) and fibronectin 6-8 repeats (FN6-8) significantly enhanced the proliferation of neural stem/progenitor cells and also obviously induced the differentiation into neurons but not oligodendrocytes. Neurite processes of neurons generated from neural progenitor cells were promoted by both EGFL and FN6-8 domains-activated microglia. Microglia triggered by EGFL and FN6-8 secreted brain-derived neurotrophic factor (BDNF) and transforming growth factor-beta (TGF-beta); interestingly, FN6-8 could activate microglia to secrete nerve growth factor in addition to BDNF and TGF-beta, but EGFL domain could not. All these data implied that the cross-talk between TN-R distinct domains EGFL/FN6-8 and microglia promoted neural stem/progenitor cell proliferation and induced their differentiation into neurons.

Herpes Simplex Virus-Thymidine Kinase–Based Suicide Gene Therapy as a “Molecular Switch Off” for Nerve Growth Factor Production In Vitro

Tissue-engineered constructs offer a new hope to patients suffering from functional impairment after nerve injury. An effort has been made to focus on delivery, regulation, and "molecular shutoff" of nerve growth factor (NGF) in tissue-engineered constructs. We have previously demonstrated that human embryonic kidney (HEK-293) cells can be genetically modified to secrete NGF at varying time points upon up regulation with Ponasterone A (PonA) both in vitro and in vivo. In the present study, HEK-293 cells that stably and inducibly produce NGF were further stably transfected with herpes simplex virus-thymidine kinase gene as a suicide gene (hNGF-EcR-293-TK) in order to shut off the NGF secretion and kill the cells upon treatment with ganciclovir (GCV). These cells following induction with PonA secreted NGF levels of 6659.2 +/- 489.4 pg/mL at day 10 postbooster dose at day 5, which was significantly higher than the control noninduced cells. The NGF secreted by these cells was bioactive as determined by a rat adrenal pheochromocytoma (PC-12) cell bioassay. Treatment of these cells with GCV significantly reduced the NGF levels to 645.3 +/- 16.2 pg/mL at day 10 and live cell numbers dropped to 7.95 x 10(3) +/- 278 compared to 2.73 x 10(5) +/- 6.1 x 10(4). GCV-treated cell media when transferred to the PC-12 cell bioassay demonstrated less than 10% cells differentiating into neurite-like extensions. We conclude that hNGF-EcR-293-TK cells can inducibly secrete bioactive NGF when treated with the inducing agent and can also be killed upon treatment with GCV. This double-gene transfection for gene expression and molecular shutoff mechanism will be a useful tool in tissue-engineered nerve constructs.

Effects of polychlorinated biphenyls (PCBs) on expression of neurotrophic factors in C6 glial cells in culture

Polychlorinated biphenyls (PCBs) are persistent environmental pollutants that affect nervous system function. Glial cells are among the first lines of defense in the nervous system and are involved in activities, including production of neurotrophic factors, which maintain an environment optimally suited for neuronal function. In this study, we investigated the effects of a commercial mixture of PCBs, Aroclor 1254 (A1254), on neurotrophic factor secretion by C6 cells in culture. C6 cells were exposed to medium containing 10 ppm A1254, 0.1% dimethyl sulfoxide (DMSO = vehicle), or normal culture medium. Glial cell line-derived neurotrophic factor (GDNF) and nerve growth factor (NGF) protein were measured by enzyme-linked immunosorbant assay. GDNF mRNA was measured by real-time RT-PCR. The role of protein kinase C (PKC) signaling in A1254 effects was investigated using bisindolylmaleimide, a PKC antagonist. Exposure to A1254 increased NGF (8.8 × 10 −5 ± 8.2 × 10 −6 pg NGF/cell) and GDNF (1.0 × 10 −4 ± 6.7 × 10 −6 pg GDNF/cell) secretion compared to DMSO treated controls (5.0 × 10 −5 ± 7.5 × 10 −6 pg NGF/cell and 6.2 × 10 −5 ± 3.1 × 10 −6 pg GDNF/cell). The effect of A1254 was long-lived, as GDNF secretion was elevated following 5 days of exposure (4.1 × 10 −5 ± 1.7 × 10 −6 pg GDNF/cell in A1254 exposed cells vs. 2.9 × 10 −5 ± 2.3 × 10 −6 pg GDNF/cell in DMSO exposed cells). GDNF mRNA expression was also elevated following exposure to A1254 (1.14 ± 0.07 gene expression units in A1254 exposed cells vs. 0.8 ± 0.07 gene expression units in DMSO exposed cells). Bisindolylmaleimide was able to block the effects of A1254 on GDNF secretion. Thus, one potential mechanism by which PCBs may alter nervous system function is via disruption of neurotrophic factor expression by glial cells. The observation that neurotrophic factor expression was increased following exposure to PCB may suggest that glial cells increase expression of neuroprotective genes following exposure to potentially damaging agents such as PCBs.

Different Roles of Dexamethasone on Transforming Growth Factor-β1–induced Fibronectin and Nerve Growth Factor Expression in Dental Pulp Cells

During pulp injury, it has been hypothesized that transforming growth factor-beta1 (TGF-beta1) is released from dentin into pulp tissue and promotes pulp tissue healing. Dexamethasone is a glucocorticoid that has been used to treat pulp injury and shown to induce differentiation of hard tissue forming cells. However, the interaction between dexamethasone and TGF-beta1 is still unknown. This study aimed to examine the effects of dexamethasone on human pulp cells in the presence of TGF-beta1. TGF-beta1 increased expression and synthesis of both fibronectin and nerve growth factor (NGF), whereas dexamethasone stimulated fibronectin synthesis but inhibited NGF expression. The application of both TGF-beta1 and dexamethasone resulted in an additional effect on fibronectin; however, dexamethasone inhibited the TGF-beta1-induced NGF expression. Dexamethasone promotes fibronectin synthesis and suppresses NGF secretion, suggesting that this reagent could be used clinically to reduce pain and promote dental pulp tissue healing.

Olfactory ensheathing cells promote migration of Schwann cells by secreted nerve growth factor

Transplantation of Schwann cells (SCs) and olfactory ensheathing cells (OECs) have emerged as very promising therapies for spinal cord repair. The important features of interaction between SCs and OECs are beginning to be appreciated, although the underlying mechanism remains unclear. In the present study, we tested the effects of OECs on SCs migration using a range of in vitro migration assays. We found that SCs migrated abundantly upon OECs monolayer, and the migration-promoting effects were identified to be due to the secreted diffusible factors in OEC-derived conditioned medium (OEC-CM). Furthermore, neutralizing nerve growth factor (NGF) in OEC-CM with NGF antibody could block this effect. Moreover, we found that NGF promotes SCs migration even on astrocyte monolayer. Taken together, these findings provide the first evidence that OECs can promote SCs migration in astrocytic environment by secreted NGF.

Patterned PLG substrates for localized DNA delivery and directed neurite extension

Tissue engineering strategies that enable nerve regeneration will require methods that can promote and direct neurite extension across the lesion. In this report, we investigate an in vitro combinatorial approach to directed neurite outgrowth using gene delivery from topographically patterned substrates, which can induce expression of neurotrophic factors to promote neurite extension and direct the extending neurites. Poly(lactide- co-glycolide) (PLG), which has been used to fabricate conduits or bridges for regeneration, was compression molded to create channels with 100, 150, and 250 μm widths. DNA complexes were immobilized to the PLG, and cells cultured on the substrate were transfected with efficiencies dependent on channel width and DNA amount. A co-culture model consisting of primary neurons and accessory cells was employed to investigate neurite outgrowth within the channels. Localized secretion of nerve growth factor (NGF) by the accessory cells promoted neuron survival and neurite extension. Neurons cultured in channels with NGF expression exhibited longer primary neurites than in the absence of channels. Neurons cultured in smaller width PLG microchannels exhibited a greater degree of directionality and less secondary sprouting than larger channels. Finally, surface immobilization allowed for the delivery of distinct plasmids from each channel, which may enable channels to be tailored for specific nerve tracts. This approach demonstrates the ability to combine gene delivery with physical guidance, and can be tailored to target specific axonal populations with varying neurotrophic factor requirements.