Interleukin-1β enhances the effect of serum deprivation on rat annular cell apoptosis

Excessive apoptosis of disc cells is believed to play an important role in intervertebral disc (IVD) degeneration. It has been shown that interleukin-1β (IL-1β) is involved in the failure of disc matrix by suppressing the synthesis of matrix components and stimulating the expression of matrix metalloproteinases. However, whether IL-1β induces disc cell apoptosis is still unclear. The objective of this study was to investigate the effect of IL-1β on the apoptosis of rat annular cells cultured with or without serum supplement. First-passage rat annular cells were cultured with 0% or 10% fetal bovine serum (FBS) supplement and stimulated with 0, 10, 20 or 50 ng/ml IL-1β for 12, 24 or 48 h. Apoptotic incidences were quantified by flow cytometry, morphologic changes in apoptotic cells were visualized by Hoechst 33258 staining and phase-contrast microscopy, and caspase-3 activity was also determined. When rat annular cells were cultured with 10% FBS supplement, no significant changes in apoptotic incidences, apoptotic morphology and caspase-3 activity were observed even when cells were stimulated with 50 ng/ml IL-1β for 48 h. In contrast, serum deprivation for 24 h led to an increase in apoptotic incidences, the number of apoptotic nuclei and caspase-3 activity, and IL-1β significantly increased the effects of serum deprivation in a dose-dependent manner. Our results indicate that IL-1β alone is not a sufficient stimulus to induce disc cell apoptosis and that in order to suppress disc cell apoptosis, improving the nutrient supply to the disc may be more effective than antagonizing the adverse effects of IL-1β.     

The involvement of p38 MAPK in transforming growth factor β1－induced apoptosis in murine hepatocytes

INTRODUCTIONApoptosis is a fundamental important biologicalprocess that is required to maintain the integrity andhomeostasis of multicenular organism[1]. It seemsthat apoptosis is a predominant type of active cendeath in the liver. Endogenous factors, such astransforming growth factor FI (TGF-gi), activin A,CD95 ligand, and tumor necrosis factor (TNF) maybe involved in induction of apoptosis in the liver[2].transforming growth factor P (TGF-P) is amember of a super-family of multifu…     

Thymosin β4 activates integrin-linked kinase and decreases endothelial progenitor cells apoptosis under serum deprivation

Thymosin β4 (Tβ4) has been suggested to regulate multiple cell signal pathways and a variety of cellular functions such as cell migration, proliferation, survival, and angiogenesis. Here, we investigated the effect of Tβ4 on endothelial progenitor cells (EPCs) apoptosis induced by serum deprivation and the corresponding signal transduction pathways involved in this process. Circulating EPCs, isolated from healthy volunteers, were cultured in the absence or presence of Tβ4 and various signal cascade inhibitors. Apoptosis was evaluated with Annexin V immunostaining and cytosolic cytochrome c expression. Incubation of EPCs with Tβ4 caused a concentration dependent increase in cell viability and proliferation activity. It also caused an inhibitory effect on EPCs apoptosis, which was abolished by PI3K inhibitors (either LY294002 or Wortmannin) or JNK MAPK inhibitor SP600125. In addition, the expression and activity of caspase-3 and -9 were decreased by Tβ4, which markedly increased the Bcl-2/Bax ratio within EPCs. Furthermore, Tβ4 was immunoprecipitated with integrin-linked kinase (ILK), accompanied by augmentation of ILK activity. Transfection of EPCs with ILK-siRNA resulted in abolishment of the activation of ILK-Akt and the ameliorative effect on apoptosis by Tβ4. Together, Tβ4 mediated inhibitory effect on EPCs apoptosis under serum deprivation can be attributed, at least in part, to ILK-Akt activation. The activation of JNK MAPK might also be involved in this process.     

The crosstalk between autophagy and apoptosis: where does this lead?

Recent advances in the understanding of the molecular processes contributing to autophagy have provided insight into the relationship between autophagy and apoptosis. In contrast to the concept of “autophagic cell death,” accumulating evidence suggests that autophagy serves a largely cytoprotective role in physiologically relevant conditions. The cytoprotective function of autophagy is mediated in many circumstances by negative modulation of apoptosis. Apoptotic signaling, in turn, serves to inhibit autophagy. While the mechanisms mediating the complex counter-regulation of apoptosis and autophagy are not yet fully understood, important points of crosstalk include the interactions between Beclin-1 and Bcl-2/Bcl-xL and between FADD and Atg5, caspase- and calpain-mediated cleavage of autophagy-related proteins, and autophagic degradation of caspases. Continued investigation of these and other means of crosstalk between apoptosis and autophagy is necessary to elucidate the mechanisms controlling the balance between survival and death both under normal conditions and in diseases including cancer.     

Vitamin D inhibition of pro-fibrotic effects of transforming growth factor β1 in lung fibroblasts and epithelial cells

The mechanisms that control fibroproliferation and matrix deposition in lung fibrosis remain unclear. We speculate that vitamin D deficiency may contribute to pulmonary fibrosis since vitamin D deficiency has been implicated in several diseases. First, we confirmed the presence of vitamin D receptors (VDRs) in cultured NIH/3T3 and lung fibroblasts. Fibroblasts transfected with a vitamin D response element–reporter construct and exposed to the active vitamin D metabolite, 1,25(OH)₂D₃, showed increased promoter activity indicating VDR functionality in these cells. Testing the effects of 1,25(OH)₂D₃ on fibroblasts treated with transforming growth factor β1 (TGFβ1), considered a driver of many fibrotic disorders, we found that 1,25(OH)₂D₃ inhibited TGFβ1-induced fibroblast proliferation in a dose-dependent fashion. 1,25(OH)₂D₃ also inhibited TGFβ1 stimulation of α-smooth muscle actin expression and polymerization and prevented the upregulation of fibronectin and collagen in TGFβ1-treated fibroblasts. Finally, we examined how 1,25(OH)₂D₃ affects epithelial–mesenchymal transformation of lung epithelial cells upon exposure to TGFβ1. We showed that the TGFβ1-induced upregulation of mesenchymal cell markers and abnormal expression of epithelial cell markers were blunted by 1,25(OH)₂D₃. These observations suggest that under TGFβ1 stimulation, 1,25(OH)₂D₃ inhibits the pro-fibrotic phenotype of lung fibroblasts and epithelial cells.     

Association between transforming growth factor β1 polymorphisms and atrial fibrillation in essential hypertensive subjects

Background  

The association of TGF β1 polymorphisms and atrial fibrillation (AF) in essential hypertensive (EH) subjects remains unknown. Methods EH subjects with AF (EH+AF+) and sinus rhythm (EH+AF-) were enrolled. The polymorphisms of +869 T → C at codon 10 and + 915 G → C at codon 25, were genotyped. The clinical characteristics including serum TGF β1 levels were detected.     

Effects of transforming growth factor β on growth of human mammary epithelial cells in culture

Summary Normal human mammary epithelial cells (HMEC) from different individual reduction mammoplasty specimens were all growth inhibited, and showed a flattened, elongated morphology in response to human recombinant transforming growth factor β1 (TGFβ). The degree of growth inhibition varied among specimens, but none of the normal HMEC maintained growth in the continued presence of TGFβ. The degree of growth inhibition also varied with cell age in vitro, cells closer to senescence being more sensitive. TGFβ sensitivity was additionally assayed in two established cell lines derived from one of the reduction mammoplasty specimens after exposure to benzo(a)pyrene. Although varying degrees of growth inhibition and morphologic changes were observed in the cell lines, both lines contained populations that maintained active growth in the presence of TGFβ. Subclones of these lines demonstrated a great plasticity in their growth response to TGFβ, with individual clones ranging from strongly growth inhibited to nearly unaffected. These results suggest that multiple factors influence the extent of TGFβ-induced growth effects on both normal and transformed mammary epithelial cells, and that some of these factors may act through epigenetic mechanisms. This work was supported by CA24844 from the National Institutes of Health, Bethesda, MD, and the Office of Energy Research, Office of Health and Environmental Research of the U.S. Department of Energy under contract DE-AC03-76SF00098.     

The response of foetal annulus fibrosus cells to growth factors: modulation of matrix synthesis by TGF-β1 and IGF-1

The annulus fibrosus of the intervertebral disc is a complex radial-ply tissue that derives initially from segmental condensations of axial mesenchyme surrounding the notochord. These mesenchymal condensations differentiate into the early annulus fibrosus during foetal development—their outer part becoming fibrous, containing collagen type I; and their inner part cartilaginous, containing type II collagen and aggrecan. With post-natal growth and ageing, there is a switch from type I to type II collagen and an increase in proteoglycan synthesis in the outer annulus. This fibrocartilaginous metaplasia appears to occur in response to compressive loading of the tissue as occurs in tendons that wrap around bony pulleys, and driven by growth factors, such as TGF-β. In this study, using high-density micromass cultures, we have assessed the response of foetal outer annulus cells to growth factor stimulation with TGF-β1 and IGF-1, growth factors known to occur within the developing disc. We qualitatively and quantitatively describe the stimulatory effects of these growth factors, both alone and in combination, on the synthesis of sulphated glycosaminoglycan, and collagen types I and II by annulus cells. We show a potential role for TGF-β1 in pushing cells towards a fibrocartilaginous phenotype, with possible complementary effects of IGF-1.     

Association between transforming growth factor β1 polymorphisms and left ventricle hypertrophy in essential hypertensive subjects

Left ventricular hypertrophy (LVH) increases the risk of cardiovascular morbid events in hypertension. TGF-β1 is involved in pathologic states such as cardiac hypertrophy and cardiac fibrosis; we thus postulate that the TGF-β1 polymorphism is related to LVH in hypertensives. Six hundred and eighty essential hypertensive patients were recruited. Biochemical variables and clinical data were obtained and the determination of LVH was performed by echocardiography. According to the presence of LVH, all subjects were divided into the LVH+ and LVH? group. DNA was obtained, and two coding region polymorphisms of the TGF-β1 gene (+869 Leu→Proat codon 10 and +915 ARG→Pro at codon 25) were analyzed by the polymerase chain reaction. The product was cleaved with the restriction endonucleases. For the polymorphisms of the +869 Leu→Pro at codon 10, there was no marked difference in the distributions of genotypes and the allele frequencies between the LVH+ and LVH? subjects. For +915 Arg→Pro at codon 25, a significant difference in the distributions of genotypes of TGF-β1 was observed. The left ventricular mass index (LVMI) in Arg-Pro genotype carriers was significantly higher than those in the Arg-Arg and Pro-Pro carriers. Multivariate analysis showed that the Arg-Pro genotype was an independent risk factor for LVH (OR 3.23, 95% CI [1.48–5.63, P = 0.002]). The codon 10 genotypes did not show a significant association to LVH. Our data revealed a genetic association of TGF-β1+915 Arg→Pro at codon 25 polymorphism with LVH in a Chinese hypertensive population.     

Effects of transforming growth factor β, tumor necrosis factor α and interferon γ on pancreatic islet β-cell responsiveness to transforming growth factor α

The insulin-producing pancreatic islet -cell, characterized by low proliferative potential, is normally not responsive to the polypeptide epidermal growth factor (EGF) or its homolog transforming growth factor (TGF-). Since EGF receptors in other tissues can be up-regulated by other growth factors and by cytokines, we have in this paper investigated whether such a -cell responsiveness to TGF-, or EGF, can be conferred by co-culture with interferon (IFN-), tumor necrosis factor (TNF-) or transforming growth factor (TGF-) in various combinations. To this end, fetal rat pancreatic islets enriched in -cells were isolated and cultured for 3 days with or without 200 pM or 20 nM TGF-. It was found that neither of these TGF- concentrations affected -cell mitogenesis, insulin content or insulin secretion. However, IFN- (1000 U/ml) evoked a modest stimulation of -cell replication, while suppressing insulin secretion and leaving the islet insulin content unaltered. TNF- (1000 U/ml), on the other hand, affected none of these parameters either alone or in any combination with TGF- or IFN-. However, when TNF- or IFN-, either alone or in combination, were combined with the cytokine interleukin-1, this resulted in islet disintegration, whereas the latter cytokine alone did not exert any gross necrotic changes evident by light microscopy. TGF- (500 pM) stimulated insulin secretion but did not influence islet insulin content or -cell mitogenesis either alone or in combination with TGF- (200 pM or 20 nM). In no instance could any mitogenic or secretory response to low or high concentrations of TGF- be conferred by IFN-, TNF- or TGF- whether used alone or in combinations. Hence, responsiveness to TGF- or EGF in the -cell obviously cannot be achieved by any of these peptides.Abbreviations EGF epidermal growth factor - IFN- interferon - TGF- transforming growth factor - TGF- transforming growth factor - TNF- tumor necrosis factor      

Immunohistochemical localization of transforming growth factor β1 and β2 in mouse testes during postnatal development

We examined age-related changes in the expression of transforming growth factor-β(1) (TGF-β(1)) and transforming growth factor-β(2) in mouse testes. The mice were assigned to three age groups: 35, 50, and 75 days old. Paraffin embedded testis sections were processed for the standard streptavidin biotin peroxidase complex immunohistochemistry method. TGF-β(1) expression increased in aging round spermatids over the time studied. There was no expression in 35-day-old Leydig cells, whereas strong expression of TGF-β(1) was observed in 50-day-old Leydig cells. Expression decreased in 75-day-old Leydig cells. TGF-β(2) expression was weak in 35- and 50-day-old mouse spermatids, but expression was greater in 75-day-old elongated spermatids. In Leydig cells, TGF-β(2) expression was strong in both 35- and 50-day-old mice, whereas the expression of TGF-β(2) was less in 75-day-old Leydig cells. Our results suggest that TGF-β(1) and TGF-β(2) may play significant roles in testicular functions and germ cell development in mice.     

Immunohistochemical localization of transforming growth factor β1 and β2 in mouse testes during postnatal development

We examined age-related changes in the expression of transforming growth factor-β₁ (TGF-β₁) and transforming growth factor-β₂ in mouse testes. The mice were assigned to three age groups: 35, 50, and 75 days old. Paraffin embedded testis sections were processed for the standard streptavidin biotin peroxidase complex immunohistochemistry method. TGF-β₁ expression increased in aging round spermatids over the time studied. There was no expression in 35-day-old Leydig cells, whereas strong expression of TGF-β₁ was observed in 50-day-old Leydig cells. Expression decreased in 75-day-old Leydig cells. TGF-β₂ expression was weak in 35- and 50-day-old mouse spermatids, but expression was greater in 75-day-old elongated spermatids. In Leydig cells, TGF-β₂ expression was strong in both 35- and 50-day-old mice, whereas the expression of TGF-β₂ was less in 75-day-old Leydig cells. Our results suggest that TGF-β₁ and TGF-β₂ may play significant roles in testicular functions and germ cell development in mice.     

The effect of β-endorphin on arginine-induced growth hormone and prolactin release

β-endorphin administration via constant infusion inhibited the release of growth hormone (GH) and augmented the release of prolactin (PRL) induced by arginine in normal female subjects. Although β-endorphin infusion also induced hyperglycemia, the increment in plasma glucose was insufficient to account for the observed suppression of arginine-initiated GH release. These studies demonstrate that β-endorphin influences, in opposed directions, the secretion of PRL and GH in women.     

Effects of transforming growth factor β1 and basic fibroblast growth factor on lipoprotein lipase activity in primary cultures of chicken (Gallus domesticus) adipocyte precursors

• 1.1. The effect of TGF-β and bFGF on lipoprotein lipase activity in chicken adipocyte precursors was investigated.
• 2.2. Lipoprotein lipase activity was reduced by up to 80% by incubation with TGF-β whereas bFGF had no effect.
• 3.3. Contrary to that found with the 3T3-L1 preadipocyte cell line it was not necessary for TGF-β to be present prior to the start of differentiation in order to be effective.
• 4.4. Incubation of adipocyte precursors with actinomycin D abolished the effect of TGF-β suggesting that synthesis of a protein effector is required.
• 5.5. These results indicate differences in responsiveness to TGF-β and bFGF between primary chicken adipocyte precursors and some preadipocyte cell lines.

     

Induction of growth cessation by acacetin via β-catenin pathway and apoptosis by apoptosis inducing factor activation in colorectal carcinoma cells

Molecular Biology Reports - Acacetin, a bioflavanoid, contains anti-inflammatory and anti-cancer activities as shown in different experimental models. However, its anticancer potential and...     

The Aarskog-Scott syndrome protein Fgd1 regulates podosome formation and extracellular matrix remodeling in transforming growth factor β-stimulated aortic endothelial cells

Podosomes are dynamic actin-rich adhesion plasma membrane microdomains endowed with extracellular matrix-degrading activities. In aortic endothelial cells, podosomes are induced by transforming growth factor β (TGF-β), but how this occurs is largely unknown. It is thought that, in endothelial cells, podosomes play a role in vessel remodeling and/or in breaching anatomical barriers. We demonstrate here that, in bovine aortic endothelial cells, that the Cdc42-specific guanine exchange factor (GEF) Fgd1 is expressed and regulated by TGF-β to induce Cdc42-dependent podosome assembly. Within 15 min of TGF-β stimulation, Fgd1, but none of the other tested Cdc42 GEFs, undergoes tyrosine phosphorylation, associates with Cdc42, and translocates to the subcortical cytoskeleton via a cortactin-dependent mechanism. Small interfering RNA-mediated Fgd1 knockdown inhibits TGF-β-induced Cdc42 activation. Fgd1 depletion also reduces podosome formation and associated matrix degradation and these defects are rescued by reexpression of Fgd1. Although overexpression of Fgd1 does not promote podosome formation per se, it enhances TGF-β-induced matrix degradation. Our results identify Fgd1 as a TGF-β-regulated GEF and, as such, the first GEF to be involved in the process of cytokine-induced podosome formation. Our findings reveal the involvement of Fgd1 in endothelial cell biology and open up new avenues to study its role in vascular pathophysiology.