Other articles by Kerstin Krieglstein on PubMed

• TGF-beta and the Regulation of Neuron Survival and Death Journal of Physiology, Paris. Jan-Mar, 2002  |   Pubmed ID: 11755780 Transforming growth factor-betas (TGF-betas) constitute a superfamily of multifunctional cytokines with important implications in morphogenesis, cell differentiation, and tissue remodeling. In the developing nervous system, TGF-beta2 and -beta3 occur in radial and astroglial cells as well as in many populations of postmitotic, differentiating neurons. TGF-beta1 is restricted to the choroid plexus and meninges. In addition to functions related to glial cell maturation and performances, TGF-beta2 and -beta3 are important regulators of neuron survival. In contrast to neurotrophic factors, as for example, neurotrophins, TGF-betas are most likely not neurotrophic by themselves. However, they can dramatically increase the potency of select neurotrophins, fibroblast growth factor-2, ciliary neurotrophic factor, and glial cell line-derived neurotrophic factor (GDNF). In the case of GDNF, we have shown that GDNF fails to promote the survival of highly purified neuron populations in vitro unless it is supplemented with TGF-beta. This also applies to the in vivo situation, where antibodies to all three TGF-beta isoforms fully prevent the trophic effect of GDNF on axotomized, target-deprived neurons. In addition to the TGF-beta isoforms -beta2 and -beta3, other members of the TGF-beta superfamily are expressed in the nervous system having important roles in embryonic patterning, cell migration, and neuronal transmitter determination. We have cloned and expressed a novel TGF-beta, named growth/differentiation factor-15 (GDF-15). GDF-15 is synthesized in the choroid plexus and released into the CSF, but also occurs in all regions investigated of the developing and adult brain. GDF-15 is a potent trophic factor for developing and 6-OHDA-lesioned midbrain dopaminergic neurons in vitro and in vivo, matching the potency of GDNF.
• Mechanisms of TGF-beta-mediated Apoptosis Cell and Tissue Research. Jan, 2002  |   Pubmed ID: 11810309 Transforming growth factor-beta (TGF-beta) is a multifunctional cytokine, whose numerous cell and tissue activities include cell-cycle control, the regulation of early development, differentiation, extracellular matrix formation, hematopoesis, angiogenesis, chemotaxis, immune functions, and the induction of apoptosis. TGF-beta-mediated growth inhibition and apoptosis can be correlated with its function as a tumor suppressor. The apoptosis-inducing capacity has been investigated in many cell types. Data from cell-culture experiments and in vivo studies argue for a pivotal role of TGF-beta-mediated apoptosis in the maintenance of B- and T-cell homeostasis. The importance of TGF-beta in the control of liver cell apoptosis and cell death of prostate epithelial cells has been confirmed in many studies. Inactivation of TGF-beta in animal models via a knockout approach or neutralizing antibodies suggests that TGF-beta-mediated apoptosis plays an important part during tissue formation and remodeling and during the phase of ontogenetic neuron death. The molecular mechanisms involved in these processes seem to involve the activation of SMAD proteins. Many studies have described an interaction of TGF-beta with other signaling cascades as exemplified by the requirement of AP1 transcription factor for the induction of apoptosis in liver cells. The aim of this review is (1) to summarize and classify data in the TGF-beta apoptosis literature with respect to the affected cell types, (2) to provide insights into the intracellular mechanisms involved in TGF-beta-mediated apoptosis, and (3) to set TGF-beta-mediated apoptosis in a physiological context.
• TGF-beta is Required for Programmed Cell Death in Interdigital Webs of the Developing Mouse Limb Mechanisms of Development. May, 2002  |   Pubmed ID: 11960699 During limb formation massive cell death in the mesenchyme of the interdigital spaces accompanies the formation of free digits. Members of the transforming growth factor beta (TGF-) superfamily were discussed to play a key role in cell-cell interactions, important in the regulation of programmed cell death (PCD). TGF-beta itself is believed to be involved in epithelial-mesenchymal interactions. Here, we demonstrate that PCD is significantly reduced in interdigital spaces of the developing limbs of Tgfbeta2-/-Tgfbeta3-/- double knockouts. The regression of interdigital webs seems to be doses-dependent as interdigital mesenchyme is at least partly reduced in Tgfbeta2-/-Tgfbeta3+/- mutants, whereas interdigital zones of Tgfbeta2-/-Tgfbeta3-/- double knockouts reveal only minimal signs of regression. We conclude that TGF- is a critical extrinsic regulator of PCD.
• The Role of Transforming Growth Factor Beta-2, Beta-3 in Mediating Apoptosis in the Murine Intestinal Mucosa Gastroenterology. May, 2002  |   Pubmed ID: 11984523 Apoptosis is especially relevant in the gastrointestinal tract because the mammalian intestinal mucosa undergoes continual epithelial regeneration. Most recently, we confirmed the proapoptotic role of endogenous transforming growth factor (TGF)-beta in the developing chick retina as well as in chick ciliary, dorsal root, and spinal motor neurons. In the present study, we determined to establish the role of TGF-beta2 and TGF-beta3 in mediating apoptosis in non-neuronal tissue by analyzing the intestinal mucosa of Tgfbeta2(+/-) and Tgfbeta3(+/-) heterozygous mice.
• TGF-beta Induces Cell Death in the Oligodendroglial Cell Line OLI-neu Glia. Oct, 2002  |   Pubmed ID: 12237847 We have shown that TGF-beta plays an important role during the period of developmental cell death in the nervous system. Immunoneutralization of TGF-beta prevents ontogenetic neuron death in vivo. Like neurons, oligodendrocytes are generated in excess and eliminated by apoptosis. It has been shown that oligodendrocyte progenitors and newly formed oligodendrocytes are especially susceptible to apoptosis. We choose the oligodendrocyte precursor cell line OLI-neu to address the question if TGF-beta could play a role for the control of oligodendrocyte proliferation and cell death. Flow cytometric analysis revealed that OLI-neu cells arrested in the G1 phase of the cell cycle underwent apoptosis in response to TGF-beta. TUNEL assays, apoptosis ELISA, and caspase assays substantiated the finding that OLI-neu cells died after TGF-beta treatment. Cell death could be inhibited by application of pan-caspase or caspase 8 and 9 inhibitors, whereas the inhibition of calpain was unaffected. Furthermore, we found a reduction of bcl-X(L) at the protein as well as at the mRNA level, while p27 was upregulated. The Smad cascade was activated while TGF-beta reduced the activity of the p42/p44 MAP kinase pathway. Together, these data show that TGF-beta induced apoptotic cell death in cells of oligodendroglial origin, whereby the signaling cascade involved the downregulation of antiapoptotic signaling such as bcl-X(L) leading to the activation of caspases.
• Transforming Growth Factor-beta Induced Cell Death in the Developing Chick Retina is Mediated Via Activation of C-jun N-terminal Kinase and Downregulation of the Anti-apoptotic Protein Bcl-X(L) Neuroscience Letters. Sep, 2002  |   Pubmed ID: 12270637 Cell death in general and especially in neuronal cells is regulated by a complex interplay between survival and death signals, generated by extracellular factors like neurotrophins and intracellular regulation mechanisms. The pleiotrophic transforming growth factor beta (TGF-beta) influences life and death decisions in cells depending on cell type and other growth factors present. It has been previously shown that TGF-beta is necessary to induce ontogenetic cell death during retinal development. In the present study, we analyzed the underlying intracellular signaling processes involved in TGF-beta mediated cell death. We established a cell culture system mimicking the situation of ontogenetic cell death in vivo with cultured retinal cells isolated from the retinae of embryonic day 7 white leghorn chick embryos. The neutralization of TGF-beta inhibits cell death of cultured retinal cells whereas exogenous application of TGF-beta is followed by enhanced apoptosis as observed by in situ cell death detection (terminal deoxynucleotidyl transferase-mediated nick end labeling) assay. TGF-beta induces the activation of c-jun N-terminal kinase in the mitogen-activated protein kinase (MAP kinase) pathway and provokes downregulation of the anti-apoptotic BCL-X(L) protein. Thus, TGF-beta influences cell death via activation of a pro-apoptotic MAP-kinase cascade accompanied by a downregulation of anti-apoptotic signals.
• Tgfbeta2 -/- Tgfbeta3 -/- Double Knockout Mice Display Severe Midline Fusion Defects and Early Embryonic Lethality Anatomy and Embryology. Dec, 2002  |   Pubmed ID: 12478370 Given all known biological activities, it is anticipated that transforming growth factors beta (TGF-betas) play important roles in many different developmental processes. As all three TGF-beta isoforms display overlapping expression patterns, deletion of one TGF-beta isoform might be compensated for by another. In the present study, targeted disruption of both Tgfbeta2 and Tgfbeta3 genes was undertaken to circumvent this problem and determine the essential roles of TGF-beta2 and TGF-beta3 in vivo. Tgfbeta2(-/-) Tgfbeta3(-/-) double knockout mice and their three-allelic Tgfbeta2(-/-) Tgfbeta3(+/-) littermates display a lack of distal parts of the rib, a lack of sternal primordia, and failure in ventral body wall closure, leading to an extrathoracic position of the heart and extrusion of the liver. In addition, abnormalities in connective tissue composition and an early embryonic lethality [around embryonic day (E) 15.5] are seen. In contrast, Tgfbeta2 (+/-) Tgfbeta3 (-/-) littermates show normal rib and sternum development, normal anterior body wall fusion, and are still alive on E18.5. TGF-beta2 is already known to play a role in skeletal and craniofacial development. The results presented here show that beyond this: (a). TGF-betas obviously play a fundamental role in midline fusion and (b). the Tgfbeta2 gene seems to play a more important role in mediating developmental processes than the Tgfbeta3 gene, since Tgfbeta2 (+/-) Tgfbeta3 (-/-) mutants - in contrast to their Tgfbeta2(-/-) Tgfbeta3 (+)(/-) littermates - do not display severe malformations.
• TGF-betas and Their Roles in the Regulation of Neuron Survival Advances in Experimental Medicine and Biology. 2002  |   Pubmed ID: 12575828 Transforming growth factor-betas (TGF-betas) are a still growing superfamily of cytokines with widespread distribution and diverse biological functions. They fall into several subfamilies including the TGF-betas 1, 2, and 3, the bone morphogenetic proteins (BMPs), the growth/differentiation factors (GDFs), activins and inhibins, and the members of the glial cell line-derived neurotrophic factor family. Following a brief description of their general roles and signaling in development, maintenance of homeostasis, and disease, we shall focus on their distribution in the CNS and their involvement in regulating neuron survival and death.
• Transforming Growth Factor Beta2 is Released from PC12 Cells Via the Regulated Pathway of Secretion Molecular and Cellular Neurosciences. Jan, 2003  |   Pubmed ID: 12595240 Transforming growth factor beta2 (TGF-beta2), a prototypic member of a large superfamily of multifunctional cytokines, is expressed by neurons and glial cells. Its subcellular compartmentalization and release from neurons, however, are largely unknown. Here we show that TGF-beta2 colocalizes with the trans-Golgi network marker TGN38 and a marker molecule for secretory granules, chromogranin B (CgB), in PC12 cells. Similarly, primary hippocampal neurons show colocalization of TGN38 and TGF-beta2. A substantial amount of endogenous as well as transfected TGF-beta2 in PC12 cells comigrates with CgB on an equilibrium gradient, suggesting costorage in secretory granules. TGF-beta biological activity peaks in identical fractions. Depolarization of PC12 cells with high potassium triggers colocalization of CgB and TGF-beta2 at the cell surface, suggesting their regulated corelease from secretory granules. High potassium also liberates biologically active TGF-beta from PC12 cells and primary neurons. Our results indicate that a substantial portion of TGF-beta2 is secreted by the regulated secretory pathway in PC12 cells and hippocampal neurons.
• Transforming Growth Factor-beta(s) Are Essential for the Development of Midbrain Dopaminergic Neurons in Vitro and in Vivo The Journal of Neuroscience : the Official Journal of the Society for Neuroscience. Jun, 2003  |   Pubmed ID: 12832542 Development of midbrain dopaminergic neurons is known to depend on inductive signals derived from the ventral midline, including Sonic hedgehog (Shh) as one of the identified molecules. Here we show that in addition to Shh, transforming growth factor (TGF)-beta is required for both induction and survival of ventrally located midbrain dopaminergic neurons. Like Shh, TGF-beta is expressed in early embryonic structures such as notochord and floor plate, as well as in the area where midbrain dopaminergic neurons are developing. Treatment of cells dissociated from the rat embryonic day (E) 12 midbrain floor with TGF-beta significantly increases the number of tyrosine hydroxylase (TH)-positive dopaminergic neurons within 24 hr. Neutralization of TGF-beta in vitro completely abolishes the induction of dopaminergic neurons. In the absence of TGF-beta, Shh cannot induce TH-positive neurons, and vice versa, neutralizing endogenous Shh abolishes the capacity of TGF-beta to induce dopaminergic neurons in vitro. Furthermore, neutralization of TGF-beta in vivo during chick E2-7 but not E4-7 resulted in a significant reduction in TH-positive neurons in the ventral midbrain floor but not in the locus coeruleus or diencephalon, which suggests that the TGF-beta is required for the induction of mesencephalic dopaminergic neurons with a critical time period at E2/E3. Furthermore, neutralization of TGF-beta between E6 and 10, a time period during maturation of mesencephalic dopaminergic neurons when no further inductive cues are required, also resulted in a significant loss of dopaminergic neurons, suggesting that TGF-beta is required for the promotion of survival of ventral midbrain dopaminergic neurons as well. Together, our results identify TGF-beta as an essential mediator for the induction and maintenance of midbrain dopaminergic neurons.
• Reduced Programmed Cell Death in the Retina and Defects in Lens and Cornea of Tgfbeta2(-/-) Tgfbeta3(-/-) Double-deficient Mice Cell and Tissue Research. Jul, 2003  |   Pubmed ID: 12838410 We have previously shown that immunoneutralization of transforming growth factor-beta (TGF-beta) in the chick embryo significantly reduces programmed cell death (PCD) in peripheral neurons, spinal cord, and retina. In order to validate these results we have begun to analyze PCD in mice with targeted ablations of the TGF-beta2 and TGF-beta3 genes. Recent analyses of mice lacking TGF-beta3 had failed to reveal an overt eye phenotype, while retinae of TGF-beta2-deficient mice showed retinal hypercellularity. We report now that eyes of Tgfbeta2/Tgfbeta3 double-deficient mice display severe alterations in the morphology of the retina, lens, and cornea. The inner neural retina-the region where TGF-beta receptor (TbetaR) I and II immunoreactivities are most prominent-is significantly thickened, and numbers of TUNEL-positive cells are significantly reduced compared to wild-type littermates. In Tgfbeta2(-/-) Tgfbeta3(-/-) and Tgfbeta2(-/-) Tgfbeta3(+/-) littermates the retina was consistently detached from the underlying pigment epithelium. Cornea, corneal stroma, and lens epithelium were significantly thinner in these mutants. In contrast, retinal morphology in Tgfbeta2(+/-) Tgfbeta3(-/-)mutant littermates resembles the situation observed in wild-type retinae except for the retinal detachment. Thus, regression in the thickness of cornea and corneal stroma seems to be TGF-beta isoform and gene dose dependent. Our results substantiate the notion based on previous analyses of chick embryos with reduced levels of endogenous TGF-beta that TGF-beta, most notably TGF-beta2, is required to mediate PCD in developing retinal cells in vivo. Moreover, our data indicate that TGF-betas play essential roles in cornea and lens development.
• Transforming Growth Factor-beta and Tumor Necrosis Factor-alpha Cooperate to Induce Apoptosis in the Oligodendroglial Cell Line OLI-neu Journal of Neuroscience Research. Aug, 2003  |   Pubmed ID: 12868066 As shown previously, transforming growth factor-beta (TGF-beta) plays an important role during the period of developmental cell death in the nervous system. As with neurons, oligodendrocytes are generated in excess and eliminated by apoptosis. The present study was aimed at investigating the possible interaction of TGF-beta with tumor necrosis factor-alpha (TNF-alpha) in the regulation of cell death in oligodendroglial precursor cells and analyzing the underlying signaling mechanisms. We show that both factors induce apoptosis independently, but cooperate when applied together. The investigation of the signaling events revealed an important role of the JNK pathway during induction of apoptosis. TGF-beta seemed to be more efficient at inducing a release in cytochrome c from mitochondria than TNF-alpha. This might be the consequence of decreased Bcl-xL levels observed in cells treated with TGF-beta but not with TNF-alpha. Both factors stimulated caspase-3 activity, which could be inhibited by caspase-8 or caspase-9 inhibitors. Therefore, we conclude that TNF-alpha and TGF-beta affect partially common pathways but also regulate different steps in the apoptotic cascade.
• Gene Structure and Evolution of Tieg3, a New Member of the Tieg Family of Proteins Gene. Jan, 2004  |   Pubmed ID: 14697507 TGF beta-inducible immediate early gene, Tieg, belongs to the superfamily of Sp1-like transcription factors containing three C(2)H(2)-zinc finger DNA binding motifs close to the C-terminus. So far, Tieg1 and Tieg2 have been identified in human and mouse. We identified Tieg3, a new member of the Tieg protein family by screening a mouse cDNA library. Tieg3 has almost all the known features of the Tieg protein family: it shares a highly conserved C(2)H(2) zinc finger DNA binding domain and is 96% identical to Tieg2 and 86% to Tieg1, respectively. In addition, the three repression domains at the N-terminus, R1, R2 and R3 are conserved in all the Tiegs. Similar to Tieg1 and Tieg2, Tieg3 mRNA is up-regulated in response to TGF beta 1 treatment and can perform the Sp1 sites mediated repression of transcription. A 4 kilobase (kb) long transcript of mouse Tieg3 can be detected using Northern-blot analysis. The gene of mouse Tieg3 contains four exons. Due to the amino acid sequence similarity, mouse Tieg2 is regarded as an orthologue of human Tieg2. However, the mouse Tieg3 gene is localized in a conserved segment on mouse chromosome 12 corresponding to human Tieg2 on chromosome 2 with the same gene order. An interesting explanation for this apparent contradiction might be a homologous recombination leading to loci exchange between the mouse Tieg3 and Tieg2.
• TIEG1 Facilitates Transforming Growth Factor-beta-mediated Apoptosis in the Oligodendroglial Cell Line OLI-neu Journal of Neuroscience Research. Feb, 2004  |   Pubmed ID: 14743447 Transforming growth factor-beta (TGF-beta) plays an important role during the period of developmental cell death in the nervous system. Using the oligodendroglial precursor cell line OLI-neu, we have previously established an in vitro system to analyze TGF-beta-mediated cell death on the molecular level. We could show that the Krüppel-like Zn-finger transcription factor TIEG1 was up-regulated after TGF-beta stimulation of OLI-neu cells and mimicked TGF-beta effects in these cells; i.e., overexpression of TIEG1 in OLI-neu cells induced apoptosis as shown by apoptosis ELISA, DNA fragmentation, and caspases-3 activation. The apoptotic pathway seemed to be initiated by repressing the expression of the antiapoptotic protein Bcl-XL. In contrast, the reporter activity of a SMAD consensus promoter was induced, whereas the promoter activity of the inhibitory SMAD7 was reduced, suggesting that SMAD-dependent TGF-beta responses, such as TGF-beta-induced apoptosis, are enhanced in the presence of TIEG1.
• Nerve Growth Factor Mediates Activation of the Smad Pathway in PC12 Cells European Journal of Biochemistry / FEBS. Mar, 2004  |   Pubmed ID: 15009204 Ligand-induced oligomerization of receptors is a key step in initiating growth factor signaling. Nevertheless, complex biological responses often require additional trans-signaling mechanisms involving two or more signaling cascades. For cells of neuronal origin, it was shown that neurotrophic effects evoked by nerve growth factor or other neurotrophins depend highly on the cooperativity with cytokines that belong to the transforming growth factor beta (TGF-beta) superfamily. We found that rat pheochromocytoma cells, which represent a model system for neuronal differentiation, are unresponsive to TGF-beta1 due to limiting levels of its receptor, TbetaRII. However, stimulation with nerve growth factor leads to activation of the Smad pathway independent of TGF-beta. In contrast to TGF-beta signaling, activation of Smad3 by nerve growth factor does not occur via phosphorylation of the C-terminal SSXS-motif, but leads to heteromeric complex formation with Smad4, nuclear translocation of Smad3 and transcriptional activation of Smad-dependent reporter genes. This response is direct and does not require de novo protein synthesis, as shown by cycloheximide treatment. This initiation of transcription is dependent on functional tyrosine kinase receptors and can be blocked by Smad7. These data provide further evidence that the Smad proteins are not exclusively activated by the classical TGF-beta triggered mechanism. The potential of NGF to activate the Smad pathway independent of TGF-beta represents an important regulatory mechanism with special relevance for the development and function of neuronal cells or of other NGF-sensitive cells, in particular those that are TGF-beta-resistant.
• GDNF Promotes Neuronal Differentiation and Dopaminergic Development of Mouse Mesencephalic Neurospheres Neuroscience Letters. May, 2004  |   Pubmed ID: 15135891 In the present study we report establishment of a neurosphere culture system derived from mouse ventral mesencephalon at embryonic day 12 and investigate effects of glial cell line-derived neurotrophic factor (GDNF) in the differentiation potential of the neurospheres. The generated neurospheres exhibit stem cell characteristics, i.e. self-renewal capacity and multipotency. Addition of exogenous GDNF resulted in neural differentiation indicated by reduced number of nestin positive cells. GDNF treatment resulted in increased numbers of beta-III-tubulin immunoreactive cells whereas glial fibrillary acidic protein immunoreactivity was not effected. Most importantly, cell numbers expressing early dopaminergic markers, Nurr1 and Ptx3, were significantly higher in GDNF-treated spheres. We conclude that GDNF promotes differentiation of mouse mesencephalic stem cells towards neuronal lineage and most notably dopaminergic development.
• TGF-beta Promotes Survival on Mesencephalic Dopaminergic Neurons in Cooperation with Shh and FGF-8 Neurobiology of Disease. Jul, 2004  |   Pubmed ID: 15193287 Impaired neuronal survival is a key event in the development of degenerative diseases, such as Parkinson's disease (PD). Here we show that transforming growth factor beta (TGF-beta) acts directly on rat E14 midbrain dopaminergic neurons in vitro, its survival-promoting effect being not mediated by BDNF, NT-3, or GDNF. Treatment with TGF-beta, sonic hedgehog (Shh), or fibroblast growth factor-8 (FGF8) significantly increased number of tyrosine hydroxylase (TH)-immunoreactive neurons after 7 days, whereas application of these factors added together further increased number of TH-positive neurons, compared to single-factor treatments. Neutralization of endogenous TGF-beta, Shh, or FGF8 significantly reduced number of dopaminergic neurons. TGF-beta treatment decreased number of apoptotic cells, having no effect on cell proliferation. Neutralization of TGF-beta in vivo during chick E6-10 resulted in reduced number of midbrain dopaminergic neurons. The results suggest that TGF-beta is required for survival of mesencephalic dopaminergic neurons acting in cooperation with Shh and FGF8.
• Factors Promoting Survival of Mesencephalic Dopaminergic Neurons Cell and Tissue Research. Oct, 2004  |   Pubmed ID: 15300492 Growth factors promoting survival of mesencephalic dopaminergic neurons are discussed in the context of their requirement during development and adulthood. The expression of growth factors should be detectable in the nigrostriatal system during critical periods of development, i.e., during the period of ontogenetic cell death and synaptogenesis and during neurite extension and neurotransmitter synthesis. Growth factors discussed include members of the family of glial-cell-line-derived neurotrophic factors (GDNF), neurotrophins, transforming growth factors beta, and low molecular compounds mimicking growth factor activities. To date, the available data support the notion that GDNF is a highly promising candidate, although GDNF-null mice lack a dopaminergic phenotype. There remains a possibility that endogenous dopaminotrophic factors remain to be discovered.
• Induction and Specification of Midbrain Dopaminergic Cells: Focus on SHH, FGF8, and TGF-beta Cell and Tissue Research. Oct, 2004  |   Pubmed ID: 15322912 Cell-fate decisions along the dorsoventral and anterior-posterior axis of the neural tube are dictated by factors from signaling and organizing centers. According to the prevailing notion, the formation of mesencephalic dopaminergic neurons is directed by diffusable signals from the notochord, floor plate, and isthmic organizer. Sonic hedgehog (Shh), secreted by the notochord and floor plate, and fibroblast growth factor (FGF) 8, secreted by the isthmus, are thought to be key molecules involved in the development of midbrain dopaminergic neurons. During the last decade, the introduction of elegant explant culture systems and the generation of transgenic and mutant mice have greatly contributed to a better understanding of the molecular signals that direct the induction and specification of midbrain dopaminergic neurons. In this context, experimental evidence has challenged the dominant roles of Shh and FGF8 in dopaminergic neuron development. Additional molecules have been identified as being required for the generation of mesencephalic dopaminergic neurons, particularly members of the transforming growth factor beta superfamily.
• The Dopaminergic Nigrostriatal System: Development, Physiology, Disease Cell and Tissue Research. Oct, 2004  |   Pubmed ID: 15340830
• Isoform-specific Role of Transforming Growth Factor-beta2 in the Regulation of Proliferation and Differentiation of Murine Adrenal Chromaffin Cells in Vivo Journal of Neuroscience Research. Nov, 2004  |   Pubmed ID: 15478122 Chromaffin cells, the neuroendocrine cells of the adrenal medulla, play an important role in molecular, cellular, and developmental neurobiology. Unlike the closely related sympathetic neurons, chromaffin cells are able to proliferate throughout their whole life span. Proliferation of chromaffin cells in vivo is thought to be regulated by the interaction of neurogenic and hormonal signals. Previous studies have shown that chromaffin cells synthesize and release transforming growth factor-betas (TGF-betas). In the present study, effects of TGF-betas on proliferation and differentiation of chromaffin cells in mouse adrenal chromaffin cells were investigated in a genetic mouse model. We observed a significant increase in the total number of tyrosine hydroxylase-positive (TH(+)) cells in Tgfbeta2(-/-) knockout mouse embryos at embryonic day (E) 18.5 compared with wild-type animals (Tgfbeta2(+/+)), but no changes in the number of TH(+) cells were observed in Tgfbeta3(-/-) mouse mutants. At E15.5, but not at E18.5, there was a marked increase in the number of proliferative cell nuclear antigen-positive chromaffin cells in Tgfbeta2(-/-) knockout embryos compared with the wild-type group. On the other hand, there was a clear decrease in the ratio of total number of phenylethanolamine-N-methyltransferase-positive cells to the total TH(+) in Tgfbeta2(-/-) mice embryos at E18.5 compared with wild-type animals. This is the first documentation of the physiological significance of the TGF-beta2, an isoform that has been suggested to play a role in the regulation of chromaffin cells proliferation and differentiation based on in vitro experiments.
• Mutation in the Gene Encoding Lysosomal Acid Phosphatase (Acp2) Causes Cerebellum and Skin Malformation in Mouse Neurogenetics. Dec, 2004  |   Pubmed ID: 15503243 We report a novel spontaneous mutation named nax in mice, which exhibit delayed hair appearance and ataxia in a homozygote state. Histological analyses of nax brain revealed an overall impairment of the cerebellar cortex. The classical cortical cytoarchitecture was disrupted, the inner granule cell layer was not obvious, the Purkinje cells were not aligned as a Purkinje cell layer, and Bergmann glias did not span the molecular layer. Furthermore, histological analyses of skin showed that the hair follicles were also abnormal. We mapped the nax locus between marker D2Mit158 and D2Mit100 within a region of 800 kb in the middle of chromosome 2 and identified a missense mutation (Gly244Glu) in Acp2, a lysosomal monoesterase. The Glu244 mutation does not affect the stability of the Acp2 transcript, however it renders the enzyme inactive. Ultrastructural analysis of nax cerebellum showed lysosomal storage bodies in nucleated cells, suggesting progressive degeneration as the underlying mechanism. Identification of Acp2 as the gene mutated in nax mice provides a valuable model system for studying the role of Acp2 in cerebellum and skin homeostasis.
• Transforming Growth Factor Beta is Required for Differentiation of Mouse Mesencephalic Progenitors into Dopaminergic Neurons in Vitro and in Vivo: Ectopic Induction in Dorsal Mesencephalon Stem Cells (Dayton, Ohio). Sep, 2006  |   Pubmed ID: 16741229 Tissue engineering is a prerequisite for cell replacement as therapeutic strategy for degenerative diseases, such as Parkinson's disease. In the present study, we investigated regional identity of mesencephalic neural progenitors and characterized their development toward ventral mesencephalic dopaminergic neurons. We show that neural progenitors from ventral and dorsal mouse embryonic day 12 mesencephalon exhibit regional identity in vitro. Treatment of ventral midbrain dissociated neurospheres with transforming growth factor beta (TGF-beta) increased the number of Nurr1- and tyrosine hydroxylase (TH)-immunoreactive cells, which can be further increased when the spheres are treated with TGF-beta in combination with sonic hedgehog (Shh) and fibroblast growth factor 8 (FGF8). TGF-beta differentiation signaling is TGF-beta receptor-mediated, involving the Smad pathway, as well as the p38 mitogen-activated protein kinase pathway. In vivo, TGF-beta2/TGF-beta3 double-knockout mouse embryos revealed significantly reduced numbers of TH labeled cells in ventral mesencephalon but not in locus coeruleus. TH reduction in Tgfbeta2(-/-)/Tgfbeta3(+/-) was higher than in Tgf-beta2(+/-)/Tgf-beta3(-/-). Most importantly, TGF-beta may ectopically induce TH-immunopositive cells in dorsal mesencephalon in vitro, in a Shh- and FGF8-independent manner. Together, the results clearly demonstrate that TGF-beta2 and TGF-beta3 are essential signals for differentiation of midbrain progenitors toward neuronal fate and dopaminergic phenotype.
• Cell Death in the Nervous System Advances in Experimental Medicine and Biology. 2006  |   Pubmed ID: 16955701
• GDNF Applied to the MPTP-lesioned Nigrostriatal System Requires TGF-beta for Its Neuroprotective Action Neurobiology of Disease. Feb, 2007  |   Pubmed ID: 17141511 GDNF is a potent neurotrophic factor for nigrostriatal dopaminergic neurons in vitro and in animal models of Parkinson's disease (PD), but has largely failed when tested in therapeutic applications in human PD. We report here that GDNF requires transforming growth factor-beta (TGF-beta) to elicit its neurotrophic activity. Lesioning the mouse nigrostriatal system with MPTP significantly upregulates striatal TGF-beta2 mRNA levels. As expected, GDNF protects against the destructive effects of MPTP, including losses of TH-ir nigral neurons, striatal dopamine and TH-ir fibers. Application of antibodies neutralizing all three TGF-beta isoforms to the MPTP-lesioned striatum abolishes the neurotrophic effect of GDNF. We show that TGF-beta antibodies are not toxic and do not interfere with retrograde transport of iodinated GDNF, suggesting that TGF-beta antibodies do not impair internalization and retrograde trafficking of GDNF. We conclude that striatal TGF-beta may be essential for permitting exogenous GDNF to act as a neuroprotective factor.
• Functional Domains of the TGF-beta-inducible Transcription Factor Tieg3 and Detection of Two Putative Nuclear Localization Signals Within the Zinc Finger DNA-binding Domain Journal of Cellular Biochemistry. Jun, 2007  |   Pubmed ID: 17252542 The recently identified TGF-beta-inducible early gene 3 (Tieg3) belongs to the gene family of Sp1/Klf-like transcription factors and is upregulated immediately after TGF-beta treatment. To explore the molecular mechanisms of Tieg3-mediated transcriptional control, GAL4-based luciferase assays were performed in order to determine regulatory domains within the Tieg3 protein. Using EGFP-fusion proteins, we monitored the intracellular localization and mapped putative nuclear localization signals (NLS). We provide evidence that the amino-terminus of Tieg3 is essential to repress the transcription and that the loss of the mSin3A interacting domain (SID) disrupts the repressive effects of Tieg3 in the oligodendroglial cell line OLI-neu. Herein we also demonstrate that the zinc finger containing DNA-binding domain (DBD) alone is able to activate the transcription of a reporter gene. Sequence analysis of the zinc finger region revealed no similarities to known activation domains. Analysis of the subcellular localization disclosed Tieg3 as a nuclear protein. Further, we identified the DBD as being essential for the nuclear localization of Tieg3. We detected two closely located putative bipartite NLS within the second and third zinc finger, which are conserved among the members of the Tieg family of proteins. Together these results may help to increase the understanding of Tieg3-mediated transcriptional control and to characterize this TGF-beta-induced Sp1/Klf-like transcription factor.
• Specificity in the Crosstalk of TGFbeta/GDNF Family Members is Determined by Distinct GFR Alpha Receptors Journal of Neurochemistry. Dec, 2007  |   Pubmed ID: 17953664 Glial cell line-derived neurotrophic factor (GDNF) and neurturin (NRTN) are neurotrophic factors for parasympathetic neurons including ciliary ganglion (CG) neurons. Recently, we have shown that survival and signaling mediated by GDNF in CG neurons essentially requires transforming growth factor beta (TGFbeta). We have provided evidence that TGFbeta regulates the availability of the glycosyl phosphatidylinositol (GPI)-anchored GDNF receptor alpha 1 (GFRalpha1) by promoting the recruitment of the receptor to the plasma membrane. We report now that in addition to GDNF, NRTN, but not persephin (PSPN) or artemin (ARTN), is able to promote survival of CG neurons. Interestingly, in contrast to GDNF, NRTN is not dependent on cooperation with TGFbeta, but efficiently promotes neuronal survival and intracellular signaling in the absence of TGFbeta. Additional treatment with TGFbeta does not further increase the NRTN response. Both NRTN and GDNF exclusively bind to and activate their cognate receptors, GFRalpha2 and GFRalpha1, respectively, as shown by the use of receptor-specific neutralizing antibodies. Immunocytochemical staining for the two receptors on the surface of CG neurons reveals that, in contrast to the effect on GFRalpha1, TGFbeta is not required for recruitment of GFRalpha2 to the plasma membrane. Moreover, binding of radioactively labeled GDNF but not NRTN is increased upon treatment of CG neurons with TGFbeta. Disruption of TGFbeta signaling does interfere with GDNF-, but not NRTN-mediated signaling and survival. We propose a model taking into account data from GFRalpha1 crystallization and ontogenetic development of the CG that may explain the differences in TGFbeta-dependence of GDNF and NRTN.
• Tieg3/Klf11 Induces Apoptosis in OLI-neu Cells and Enhances the TGF-beta Signaling Pathway by Transcriptional Repression of Smad7 Journal of Cellular Biochemistry. Jun, 2008  |   Pubmed ID: 18189266 TGF-beta signaling is indispensible for development of the nervous system since it regulates ontogenetic cell death. The recently identified TGF-beta-inducible zinc finger protein Tieg3/Klf11 belongs to the family of Sp1/Klf-like transcription factors and shares all structural and functional features with other Tieg proteins. Using the established TGF-beta-responsive oligodendroglial cell line OLI-neu, we analyzed the role of Tieg3/Klf11 in TGF-beta signaling. In this report, we show that Tieg3/Klf 11 mimics TGF-beta effects by inducing apoptotic cell death accompanied by activation of caspase-3. Moreover, we demonstrate that Tieg3/Klf11 enhances TGF-beta signaling by transcriptional repression of the inhibitory Smad7 and, thereby, disrupts the negative feedback loop of the TGF-beta signaling pathway. Loss of the N-terminal repression domains of Tieg3/Klf11 abrogates the pro-apoptotic nature of this transcription factor and abolishes the enhancement of Smad-mediated TGF-beta responses. In conclusion, we provide evidence that the recently identified transcription factor Tieg3/Klf11 is a downstream mediator of TGF-beta-induced apoptosis in the oligodendroglial cell line OLI-neu. Since other signaling molecules are able to initiate transcription of members of the Tieg family, the ability of Tieg3/Klf11 to modulate TGF-beta signaling by transcriptional inhibition of Smad7 might be an important clue for the understanding of the crosstalk between different signaling pathways.
• Transforming Growth Factor Beta Cooperates with Persephin for Dopaminergic Phenotype Induction Stem Cells (Dayton, Ohio). Jul, 2008  |   Pubmed ID: 18420832 The aim of the present study was to investigate the putative cooperative effects of transforming growth factor beta (TGF-beta) and glial cell line-derived neurotrophic factor (GDNF) family ligands in the differentiation of midbrain progenitors toward a dopaminergic phenotype. Therefore, a mouse midbrain embryonic day (E) 12 neurospheres culture was used as an experimental model. We show that neurturin and persephin (PSPN), but not GDNF, are capable of transient induction of dopaminergic neurons in vitro. This process, however, requires the presence of endogenous TGF-beta. In contrast, after 8 days in vitro GDNF rescued the TGF-beta neutralization-dependent loss of the TH-positive cells. In vivo, at E14.5, no apparent phenotype concerning dopaminergic neurons was observed in Tgf-beta2(-/-)/gdnf(-/-) double mutant mice. In vitro, combined TGF-beta/PSPN treatment achieved a yield of approximately 20% TH-positive cells that were less vulnerable against 1-methyl-4-phenyl pyridinium ion toxicity. The underlying TGF-beta/PSPN differentiation signaling is receptor-mediated, involving p38 mitogen-activated protein kinase and phosphatidylinositol 3-kinase pathways. These results indicate that phenotype induction and survival of fully differentiated neurons are accomplished through distinct pathways and individual factor requirement. TGF-beta is required for the induction of dopaminergic neurons, whereas GDNF is required for regulating and/or maintaining a differentiated neuronal phenotype. Moreover, this study suggests that the combination of TGF-beta with PSPN is a potent inductive cocktail for the generation of dopaminergic neurons that should be considered in tissue engineering and cell replacement therapies for Parkinson's disease.
• Loss of Transforming Growth Factor-beta 2 Leads to Impairment of Central Synapse Function Neural Development. 2008  |   Pubmed ID: 18854036 The formation of functional synapses is a crucial event in neuronal network formation, and with regard to regulation of breathing it is essential for life. Members of the transforming growth factor-beta (TGF-beta) superfamily act as intercellular signaling molecules during synaptogenesis of the neuromuscular junction of Drosophila and are involved in synaptic function of sensory neurons of Aplysia.
• TGF-beta Superfamily Members, ActivinA and TGF-beta1, Induce Apoptosis in Oligodendrocytes by Different Pathways Cell and Tissue Research. Dec, 2008  |   Pubmed ID: 19002501 Activins and transforming growth factor (TGF)-betas, members of the TGF-beta superfamily, affect numerous physiological processes, including apoptosis, in a variety of organs and tissues. Apoptotic functions of TGF-betas, in contrast to those of the activins, are well documented in the developing and adult nervous system. TGF-betas operate in a context-dependent manner and cooperate with other cytokines in the regulation of apoptosis. In this study, we show, for the first time, an apoptotic function of ActivinA in the nervous system, i.e. in oligodendroglial progenitor cells. Using the oligodendroglial cell line OLI-neu, we show that ActivinA acts autonomously, without cooperating with TGF-beta. In contrast to the mechanism of TGF-beta-mediated apoptosis involving Bcl-xl down-regulation, Bcl-xl in ActivinA-induced apoptosis is classically sequestered by the BH3-only protein Puma. Puma expression is controlled by the transcription factor p53 as demonstrated by experiments with the p53 inhibitor Pifithrin-alpha. Furthermore, in the apoptotic TGF-beta pathway, caspase-3 is activated, whereas in the apoptotic ActivinA pathway, apoptosis-inducing factor is released to trigger DNA fragmentation. These data suggest that TGF-beta and ActivinA induce apoptosis in oligodendrocytes by different apoptotic pathways.
• In Vivo Requirement of TGF-beta/GDNF Cooperativity in Mouse Development: Focus on the Neurotrophic Hypothesis International Journal of Developmental Neuroscience : the Official Journal of the International Society for Developmental Neuroscience. Feb, 2009  |   Pubmed ID: 18824086 Neurotrophic factors are well-recognized extracellular signaling molecules that regulate neuron development including neurite growth, survival and maturation of neuronal phenotypes in the central and peripheral nervous system. Previous studies have suggested that TGF-beta plays a key role in the regulation of neuron survival and death and potentiates the neurotrophic activity of several neurotrophic factors, most strikingly of GDNF. To test the physiological relevance of this finding, TGF-beta2/GDNF double mutant (d-ko) mice were generated. Double mutant mice die at birth like single mutants due to kidney agenesis (GDNF-/-) and congential cyanosis (TGF-beta2-/-), respectively. To test for the in vivo relevance of TGF-beta2/GDNF cooperativity to regulate neuron survival, mesencephalic dopaminergic neurons, lumbar motoneurons, as well as neurons of the lumbar dorsal root ganglion and the superior cervical ganglion were investigated. No loss of mesencephalic dopaminergic neurons was observed in double mutant mice at E18.5. A partial reduction in neuron numbers was observed in lumbar motoneurons, sensory and sympathetic neurons in GDNF single mutants, which was further reduced in TGF-beta2/GDNF double mutant mice at E18.5. However, TGF-beta2 single mutant mice showed no loss of neurons. These data point towards a cooperative role of TGF-beta2 and GDNF with regard to promotion of survival within the peripheral motor and sensory systems investigated.
• Involvement of Fractin in TGF-beta-induced Apoptosis in Oligodendroglial Progenitor Cells Glia. Nov, 2009  |   Pubmed ID: 19330858 Transforming growth factor-beta (TGF-beta) induces apoptotic cell death during the development of the nervous system. We recently identified that TGF-beta induced apoptosis in oligodendroglial progenitor cells (primary cells as well as oligodendroglial cell line OLI-neu) is characterized by down-regulation of Bcl-xl. In this report, we now focused on mechanisms that mediate TGF-beta dependent Bcl-xl down-regulation in oligodendroglial cells. We showed that the caspase-specific cleavage product Fractin is produced in oligodendroglial cells during TGF-beta-mediated apoptosis, which represents an early event of the cascade. Cleavage of actin into Fractin was dependent on functional actin and caspases, and occurred simultaneously with a Fractin-Bcl-xl-interaction. This Fractin-Bcl-xl interaction indicated a connection between Bcl-xl down-regulation and Fractin appearance, since Bcl-xl regulation was also dependent on caspases and functional actin, and an overexpression of Fractin induced a Bcl-xl protein down-regulation. Further analysis of Fractin-Bcl-xl interaction in other culture systems confirmed these data. In conclusion, we show that Fractin is not only an apoptotic marker, but has indeed a functional role in apoptotic signaling in oligodendrocytes.
• Differentiation of Embryonic Stem Cells Current Protocols in Neuroscience / Editorial Board, Jacqueline N. Crawley ... [et Al.]. Apr, 2009  |   Pubmed ID: 19340810 Mouse embryonic stem (ES) cells are derived from mouse blastocyst and are able to generate all embryonic tissues in vitro. This propensity of ES cells has acquired considerable attention in recent years due to the promising potential for future cell replacement-based therapies. Therefore, it is of fundamental interest to establish protocols that allow the differentiation of ES cells into specific cell types. In recent years, several such differentiation procedures have been described for mouse and human embryonic stem cells. This unit describes a simple procedure that promotes the neuronal differentiation of mouse embryonic stem cells and yields a high proportion of midbrain dopaminergic neurons. Furthermore, this procedure permits the isolation of neural stem cell lines from mouse ES cells.
• TGF-beta in Dopamine Neuron Development, Maintenance and Neuroprotection Advances in Experimental Medicine and Biology. 2009  |   Pubmed ID: 19731553 ransforming growth factor betas (TGF-betas) are multifunctional cytokines with widespread distribution.In the nervous system the biological effects of TGF-beta cover regulation of proliferation, migration, differentiation, survival and death. Specifically, the effects of TGF-(3 on mesencephalic DAergic neurons extend from induction and specification of the dopaminergic phenotype via promotion of survival to neuroprotection in animal models of parkinsonism. Experimental in vitro and in vivo models have contributed to a better understanding of the putative mechanisms underlying the effects of TGF-beta on DAergic neurons and unravelled synergisms between members of the TGF-beta superfamily. In this chapter, we will review the literature available with focus on TGF-beta proper and glial cell-line-derived neurotrophic factor (GDNF).
• Transforming Growth Factor Beta Promotes Neuronal Cell Fate of Mouse Cortical and Hippocampal Progenitors in Vitro and in Vivo: Identification of Nedd9 As an Essential Signaling Component Cerebral Cortex (New York, N.Y. : 1991). Mar, 2010  |   Pubmed ID: 19587023 Transforming Growth Factor beta (Tgfbeta) and associated signaling effectors are expressed in the forebrain, but little is known about the role of this multifunctional cytokine during forebrain development. Using hippocampal and cortical primary cell cultures of developing mouse brains, this study identified Tgfbeta-regulated genes not only associated with cell cycle exit of progenitors but also with adoption of neuronal cell fate. Accordingly, we observed not only an antimitotic effect of Tgfbeta on progenitors but also an increased expression of neuronal markers in Tgfbeta treated cultures. This effect was dependent upon Smad4. Furthermore, in vivo loss-of-function analyses using Tgfbeta2(-/-)/Tgfbeta3(-/-) double mutant mice showed the opposite effect of increased cell proliferation and fewer neurons in the cerebral cortex and hippocampus. Gata2, Runx1, and Nedd9 were candidate genes regulated by Tgfbeta and known to be involved in developmental processes of neuronal progenitors. Using siRNA-mediated knockdown, we identified Nedd9 as an essential signaling component for the Tgfbeta-dependent increase in neuronal cell fate. Expression of this scaffolding protein, which is mainly described as a signaling molecule of the beta1-integrin pathway, was not only induced after Tgfbeta treatment but was also associated with morphological changes of the Nestin-positive progenitor pool observed upon exposure to Tgfbeta.
• Tieg1/Klf10 is Upregulated by NGF and Attenuates Cell Cycle Progression in the Pheochromocytoma Cell Line PC12 Journal of Neuroscience Research. Jul, 2010  |   Pubmed ID: 20155803 The transcription factor Tieg1/Klf10 belongs to a family of Sp1/Klf proteins that have been shown to play important roles during development and maintenance of various tissues and cell types. Upregulation of Tieg1/Klf10 has been reported for TGF-beta, BMP2, BMP4, ActivinA and GDNF as members of the TGF-beta superfamily. Moreover, estrogen, the cytostatic drugs homoharringtonine and velcade as well as nitric oxide are also able to trigger Tieg1/Klf10 transcription. Recent studies suggest a role for members of the neurotrophin family in regulating Tieg1/Klf10 transcriptional upregulation. Using semi-quantitative RT-PCR and immunoblotting, we present data describing that nerve growth factor (NGF) regulates the expression of Tieg1/Klf10 in the pheochromocytoma cell line PC12 in a TrkA-dependent manner. Moreover, we provide evidence for the existence of NGF-responsive elements in the 5'-regulatory region of Tieg1/Klf10 that contain binding sites for the transcription factors Sp1 and CREB. After treatment with NGF PC12 cells exit the cell cycle and start to differentiate towards a neuron-like phenotype indicated by neurite outgrowth. Using flow cytometry and differentiation assays we demonstrate that Tieg1/Klf10 reduces cell cycle progression in PC12 cells but fails to promote their terminal differentiation. Together, our results identify Tieg1/Klf10 as a new NGF target gene and substantiate its anti-proliferative function in the NGF signaling pathway in PC12 cells.
• TGF-beta 1 Enhances Neurite Outgrowth Via Regulation of Proteasome Function and EFABP Neurobiology of Disease. Jun, 2010  |   Pubmed ID: 20211260 Malfunction of the ubiquitin-proteasome system has been implicated as a causal factor in the pathogenesis of aggregation-related disorders, e.g. Parkinson's disease. We show here that Transforming growth factor-beta 1 (TGF-beta), a multifunctional cytokine and trophic factor for dopaminergic (DAergic) neurons modulates proteasome function in primary midbrain neurons. TGF-beta differentially inhibited proteasomal subactivities with a most pronounced time-dependent inhibition of the peptidyl-glutamyl peptide hydrolyzing-like and chymotrypsin-like subactivity. Regulation of proteasomal activity could be specifically quantified in the DAergic subpopulation. Protein blot analysis revealed an accumulation of ubiquitinated proteins after TGF-beta treatment. The identity of these enriched proteins was further analyzed by 2D-gel electrophoresis and mass spectrometry. We found epidermal fatty acid binding protein (EFABP) to be strongly increased and ubiquitinated after TGF-beta treatment and confirmed this finding by co-immunoprecipitation. While application of TGF-beta increased neurite regeneration in a scratch lesion model, downregulation of EFABP by siRNA significantly decreased this effect. We thus postulate that a differential regulation of proteasomal function, as demonstrated for TGF-beta, can result in an enrichment of proteins, such as EFABP, that mediate physiological functions, such as neurite regeneration.
• Aged Tgfbeta2/Gdnf Double-heterozygous Mice Show No Morphological and Functional Alterations in the Nigrostriatal System Journal of Neural Transmission (Vienna, Austria : 1996). Jun, 2010  |   Pubmed ID: 20458508 Loss of dopaminergic neurons in the substantia nigra pars compacta and the resulting decrease in striatal dopamine levels are the hallmarks of Parkinson's disease. Tgfbeta and Gdnf have been identified as neurotrophic factors for dopaminergic midbrain neurons in vivo and in vitro. Haploinsufficiency for either Tgfbeta or Gdnf led to dopaminergic deficits. In this study we therefore analyzed the nigrostriatal system of aged Tgfbeta2 (+/-)/Gdnf (+/-) double-heterozygous mice. Unexpectedly, we found no morphological changes in the nigrostriatal system as compared with age-matched wild-type mice. There were no significant differences in the number of TH-positive midbrain neurons and no changes in the optical density of TH immunoreactivity in striata of Tgfbeta2 (+/-)/Gdnf (+/-) double-heterozygous mice. Moreover, we found no significant differences in the striatal levels of dopamine and its metabolites dihydroxyphenylacetic acid and homovanillic acid. Our results indicate that a combined haploinsufficiency for Tgfbeta2 and Gdnf has no impact on the function and the survival of midbrain DA neurons under normal aging conditions.
• Thapsigargin-induced Ca2+ Increase Inhibits TGFβ1-mediated Smad2 Transcriptional Responses Via Ca2+/calmodulin-dependent Protein Kinase II Journal of Cellular Biochemistry. Dec, 2010  |   Pubmed ID: 20717921 Transforming growth factor β (TGFβ) signalling plays important roles in a variety of tissues and cell types. Impaired TGFβ signalling contributes to several pathologies, including cancer, fibrosis as well as neurodegenerative diseases. TGFβ receptor type I-mediated phosphorylation of Smad2, the formation of the Smad2-Smad4 complex and translocation to the nucleus are critical steps of the TGFβ signalling pathway. Here, we demonstrate that thapsigargin-mediated increase of intracellular Ca(2+) concentrations inhibited TGFβ1-induced Smad2 transcriptional activity in the oligodendroglial cell line OLI-neu. We provide evidence that thapsigargin treatment dramatically reduced the nuclear translocation of Smad2 after TGFβ1 treatment but had no effect on its phosphorylation at Ser465/467. Moreover, using Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) inhibitors and a constitutively active CaMKII mutant, we provide evidence that the observed inhibition of TGFβ signalling in OLI-neu cells was strongly dependent on Ca(2+)-mediated CaMKII activation. In summary, this study clearly shows that the TGFβ1-induced Smad2 nuclear translocation is negatively regulated by intracellular Ca(2+) in OLI-neu cells and that increased intracellular Ca(2+) concentrations block Smad2-mediated transcription of TGFβ target genes. These results underline the importance of intracellular Ca(2+) for the regulation of TGFβ signalling.
• Lack of the Endosomal SNAREs Vti1a and Vti1b Led to Significant Impairments in Neuronal Development Proceedings of the National Academy of Sciences of the United States of America. Feb, 2011  |   Pubmed ID: 21262811 Fusion between membranes is mediated by specific SNARE complexes. Here we report that fibroblasts survive the absence of the trans-Golgi network/early endosomal SNARE vti1a and the late endosomal SNARE vti1b with intact organelle morphology and minor trafficking defects. Because vti1a and vti1b are the only members of their SNARE subclass and the yeast homolog Vti1p is essential for cell survival, these data suggest that more distantly related SNAREs acquired the ability to function in endosomal traffic during evolution. However, absence of vti1a and vti1b resulted in perinatal lethality. Major axon tracts were missing, reduced in size, or misrouted in Vti1a(-/-) Vti1b(-/-) embryos. Progressive neurodegeneration was observed in most Vti1a(-/-) Vti1b(-/-) peripheral ganglia. Neurons were reduced by more than 95% in Vti1a(-/-) Vti1b(-/-) dorsal root and geniculate ganglia at embryonic day 18.5. These data suggest that special demands for endosomal membrane traffic could not be met in Vti1a(-/-) Vti1b(-/-) neurons. Vti1a(-/-) and Vti1b(-/-) single deficient mice were viable without these neuronal defects, indicating that they can substitute for each other in these processes.
• Fibrosis and Evidence for Epithelial-mesenchymal Transition in the Kidneys of Patients with Staghorn Calculi BJU International. Oct, 2011  |   Pubmed ID: 21410631 • To quantify fibrotic lesions in renal tissues obtained from patients with large calculi and to evaluate association with renal function. • Presence of epithelial-mesenchymal transition (EMT) in stone-containing renal tissues was investigated.
• Sim1 is a Novel Regulator in the Differentiation of Mouse Dorsal Raphe Serotonergic Neurons PloS One. 2011  |   Pubmed ID: 21541283 Mesencephalic dopaminergic neurons (mDA) and serotonergic (5-HT) neurons are clinically important ventral neuronal populations. Degeneration of mDA is associated with Parkinson's disease; defects in the serotonergic system are related to depression, obsessive-compulsive disorder, and schizophrenia. Although these neuronal subpopulations reveal positional and developmental relationships, the developmental cascades that govern specification and differentiation of mDA or 5-HT neurons reveal missing determinants and are not yet understood.
• More Than Being Protective: Functional Roles for TGF-β/activin Signaling Pathways at Central Synapses Trends in Neurosciences. Aug, 2011  |   Pubmed ID: 21742388 It is becoming increasingly clear that members of the transforming growth factor-β (TGF-β) family have roles in the central nervous system that extend beyond their well-established roles as neurotrophic and neuroprotective factors. Recent findings have indicated that the TGF-β signaling pathways are involved in the modulation of both excitatory and inhibitory synaptic transmission in the adult mammalian brain. In this review, we discuss how TGF-β, bone morphogenetic protein and activin signaling at central synapses modulate synaptic plasticity, cognition and affective behavior. We also discuss the implications of these findings for the molecular understanding and potential treatment of neuropsychiatric diseases, such as anxiety, depression and other neurological disorders.
• Neuregulin 1 Type III/ErbB Signaling is Crucial for Schwann Cell Colonization of Sympathetic Axons PloS One. 2011  |   Pubmed ID: 22194888 Analysis of Schwann cell (SC) development has been hampered by the lack of growing axons in many commonly used in vitro assays. As a consequence, the molecular signals and cellular dynamics of SC development along peripheral axons are still only poorly understood. Here we use a superior cervical ganglion (SCG) explant assay, in which axons elongate after treatment with nerve growth factor (NGF). Migration as well as proliferation and apoptosis of endogenous SCG-derived SCs along sympathetic axons were studied in these cultures using pharmacological interference and time-lapse imaging. Inhibition of ErbB receptor tyrosine kinases leads to reduced SC proliferation, increased apoptosis and thereby severely interfered with SC migration to distal axonal sections and colonization of axons. Furthermore we demonstrate that SC colonization of axons is also strongly impaired in a specific null mutant of an ErbB receptor ligand, Neuregulin 1 (NRG1) type III. Taken together, using a novel SC development assay, we demonstrate that NRG1 type III serves as a critical axonal signal for glial ErbB receptors that drives SC development along sympathetic axons.
• Klf10 and Klf11 As Mediators of TGF-beta Superfamily Signaling Cell and Tissue Research. Jan, 2012  |   Pubmed ID: 21574058 Klf10 and Klf11 belong to the family of Sp1/Krüppel-like zinc finger transcription factors that play important roles in a variety of cell types and tissues. Although Klf10 and Klf11 were initially introduced as transforming growth factor-beta (TGF-beta)-inducible genes, several studies have described their upregulation by a plethora of growth factors, cytokines and hormones. Here, we review the current knowledge of the inductive cues for Klf10 and Klf11 and focus on their transcriptional regulation by members of the TGF-beta superfamily. We further summarize their involvement in the regulation of the TGF-beta signaling pathway and discuss their possible role as molecules mediating crosstalk between various signaling pathways. Finally, we provide an overview of the pro-apoptotic and anti-proliferative functions of Klf10 and Klf11.
• IL6 Protects MN9D Cells and Midbrain Dopaminergic Neurons from MPP(+)-Induced Neurodegeneration Neuromolecular Medicine. Jul, 2012  |   Pubmed ID: 22772723 The degeneration of midbrain dopaminergic (mDA) neurons is the hallmark of Parkinson's disease (PD), and several in vivo and in vitro models have been established to resemble the processes occurring during disease progression. One of the most commonly used disease models for PD is the toxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), which selectively kills mDA neurons when applied systemically. In vivo, MPTP intoxication is accompanied by a strong microglia response which is characterised by the release of inflammatory molecules such as tumour necrosis factor alpha (TNF-alpha) and interleukin-6 (IL6) that are believed to further drive inflammation-mediated degeneration of mDA neurons. Here, we addressed the question whether primary ventral mDA neurons and MN9D cells release cytokines in vitro and how these cytokine profiles change after treatment with MPP(+). Our results demonstrate that both culture models show different cytokine profiles under control conditions indicating that comparisons between both models should be made very carefully. Moreover, MN9D cells released high levels of IL6 and IP10/CXCL10, both of which were down regulated after treatment with MPP(+). MN9D-derived IL6 seems to be important for MN9D survival since neutralisation of endogenous IL6 resulted in degeneration of MN9D cells. Moreover, recombinant IL6 was able to rescue MN9D cells and primary mDA neuron cultures from MPP(+)-induced neurotoxicity, underlining the neuroprotective properties of IL6.
• Schwann Cells Migrate Along Axons in the Absence of GDNF Signaling BMC Neuroscience. 2012  |   Pubmed ID: 22863354 During development neural crest derived Schwann Cell (SC) precursors migrate to nerve trunks and populate nascent nerves. Axonal ensheathment by SC is a prerequisite for normal nerve function and the integrity of myelinated as well as nonmyelinated axons. To provide adequate support functions, SC colonize entire nerves. One important prerequisite for this is their migration into distal axonal regions.
• TGFβ Signalling Plays an Important Role in IL4-induced Alternative Activation of Microglia Journal of Neuroinflammation. 2012  |   Pubmed ID: 22947253 ABSTRACT:
• Endogenous Transforming Growth Factor-beta Promotes Quiescence of Primary Microglia in Vitro Glia. Oct, 2012  |   Pubmed ID: 23065670 Microglia are the immune cells of the central nervous system (CNS) and play important roles under physiological and pathophysiological conditions. Activation of microglia has been reported for a variety of CNS diseases and is believed to be involved in inflammation-mediated neurodegeneration. Loss of TGFβ1 results in increased microgliosis and neurodegeneration in mice which indicates that TGFβ1 is an important regulator of microglial functions in vivo. Here, we addressed the role of endogenous TGFβ signaling for microglia in vitro. We clearly demonstrate active TGFβ signaling in primary microglia and further introduce Klf10 as a new TGFβ target gene in microglia. Moreover, we provide evidence that microglia express and release TGFβ1 that acts in an autocrine manner to activate microglial TGFβ/Smad signaling in vitro. Using microarrays, we identified TGFβ-regulated genes in microglia that are involved in TGFβ1 processing, its extracellular storage as well as activation of latent TGFβ. Finally, we demonstrate that pharmacological inhibition of microglial TGFβ signaling resulted in upregulation of the proinflammatory markers IL6 and iNOS and downregulation of the alternative activation markers Arg1 and Ym1 in vitro. Together, these data clearly show that endogenous TGFβ1 and autocrine TGFβ signaling is important for microglial quiescence in vitro and further suggest the upregulation of TGFβ1 in neurodegenerative diseases as a mechanism to regulate microglia functions and silence neuroinflammation. © 2012 Wiley Periodicals, Inc.