Redox status regulates autophagy in thymic stromal cells and promotes T cell tolerance.

Thymic stromal cells (TSCs) are critical regulators of T cell tolerance, but their basic biology has remained under-characterized because they are relatively rare and difficult to isolate. Recent work has revealed that constitutive autophagy in TSCs is required for self-antigen presentation and central T cell tolerance induction; however, the mechanisms regulating constitutive autophagy in TSCs are not well understood. Hydrogen peroxide has been shown to increase autophagy flux in other tissues, and we previously identified conspicuously low expression of the hydrogen peroxide-quenching enzyme catalase in TSCs. We investigated whether the redox status of TSCs established by low catalase expression regulates their basal autophagy levels and their capacity to impose central T cell tolerance. Transgenic overexpression of catalase diminished autophagy in TSCs and impaired thymocyte clonal deletion, concomitant with increased frequencies of spontaneous lymphocytic infiltrates in lung and liver and of serum antinuclear antigen reactivity. Effects on clonal deletion and autoimmune indicators were diminished in catalase transgenic mice when autophagy was rescued by expression of the Becn1F121A/F121A knock-in allele. These results suggest a metabolic mechanism by which the redox status of TSCs may regulate central T cell tolerance.

Delta-like 4-Derived Notch Signals Differentially Regulate Thymic Generation of Skin-Homing CCR10+NK1.1+ Innate Lymphoid Cells at Neonatal and Adult Stages.

The thymus is a primary lymphoid organ for T cell development. Increasing evidence found that the thymus is also an important site for development of innate lymphoid cells (ILCs). ILCs generated in thymi acquire unique homing properties that direct their localization into barrier tissues such as the skin and intestine, where they help local homeostasis. Mechanisms underlying the developmental programming of unique tissue-homing properties of ILCs are poorly understood. We report in this article that thymic stroma-derived Notch signaling is differentially involved in thymic generation of a population of NK1.1+ group 1 ILCs (ILC1s) with the CCR10+ skin-homing property in adult and neonatal mice. We found that thymic generation of CCR10+NK1.1+ ILC1s is increased in T cell-deficient mice at adult, but not neonatal, stages, supporting the notion that a large number of developing T cells interfere with signals required for generation of CCR10+NK1.1+ ILC1s. In an in vitro differentiation assay, increasing Notch signals promotes generation of CCR10+NK1.1+ ILC1s from hematopoietic progenitors. Knockout of the Notch ligand Delta-like 4 in thymic stroma impairs generation of CCR10+NK1.1+ ILC1s in adult thymi, but development of CCR10+NK1.1+ ILC1s in neonatal thymi is less dependent on Delta-like 4-derived Notch signals. Mechanistically, the Notch signaling is required for proper expression of the IL-7R CD127 on thymic NK1.1+ ILC1s, and deficiency of CD127 also impairs thymic generation of CCR10+NK1.1+ ILC1s at adult, but not perinatal, stages. Our findings advanced understanding of regulatory mechanisms of thymic innate lymphocyte development.

Artery buckling stimulates cell proliferation and NF-κB signaling.

Tortuous carotid arteries are often seen in aged populations and are associated with atherosclerosis, but the underlying mechanisms to explain this preference are unclear. Artery buckling has been suggested as one potential mechanism for the development of tortuous arteries. The objective of this study, accordingly, was to determine the effect of buckling on cell proliferation and associated NF-κB activation in arteries. We developed a technique to generate buckling in porcine carotid arteries using long artery segments in organ culture without changing the pressure, flow rate, and axial stretch ratio. Using this technique, we examined the effect of buckling on arterial wall remodeling in 4-day organ culture under normal and hypertensive pressures. Cell proliferation, NF-κB p65, IκB-α, ERK1/2, and caspase-3 were detected using immunohistochemistry staining and immunoblot analysis. Our results showed that cell proliferation was elevated 5.8-fold in the buckling group under hypertensive pressure (n = 7, P < 0.01) with higher levels of NF-κB nuclear translocation and IκB-α degradation (P < 0.05 for both). Greater numbers of proliferating cells were observed on the inner curve side of the buckled arteries compared with the outer curve side (P < 0.01). NF-κB colocalized with proliferative nuclei. Computational simulations using a fluid-structure interaction model showed reduced wall stress on the inner side of buckled arteries and elevated wall stress on the outer side. We conclude that arterial buckling promotes site-specific wall remodeling with increased cell proliferation and NF-κB activation. These findings shed light on the biomechanical and molecular mechanisms of the pathogenesis of atherosclerosis in tortuous arteries.

Elevated serum high-sensitivity C-reactive protein levels in fibromyalgia syndrome patients correlate with body mass index, interleukin-6, interleukin-8, erythrocyte sedimentation rate.

The levels of several inflammatory cytokines are abnormal in many patients with the fibromyalgia syndrome (FMS) and may play a role in its pathogenesis. The inflammatory marker C-reactive protein (CRP) is associated with the disease activity in patients with inflammatory rheumatic diseases, but its role in FMS is unknown. We undertook this study to determine whether high-sensitivity CRP (hsCRP) is elevated in FMS and whether its levels relate to key biologic or clinical measures. One hundred and five patients with FMS (1990 ACR criteria) and 61 healthy normal controls (HNC) at a ratio of 2:1 were recruited. The serum concentrations of hsCRP, interleukin-8 (IL-8), and interleukin-6 (IL-6) were assessed using enzyme-linked immunosorbent assays. The hsCRP levels were marginally higher in FMS than in HNC (p = 0.06) and its abnormality rate (>1.5 SD above the HNC mean) was significantly higher in FMS (25 %) compared with HNC (6.8 %) (p = 0.03). Serum IL-8 levels, IL-6 levels, and erythrocyte sedimentation rate (ESR) in FMS did not differ from those in HNC. Body mass index (BMI), ESR, IL-8, and IL-6 levels correlated with hsCRP levels in FMS. No associations were found between hsCRP and age, gender, ethnicity, or other clinical measures. Serum CRP levels were higher in FMS and significantly correlated with BMI, ESR, IL-8, and IL-6 levels, suggesting that inflammation may contribute to the symptoms in some FMS patients, particularly those who are obese. Weight loss and therapies directed against inflammation may be useful in the management of FMS patients with elevated hsCRP.

A brain-derived neurotrophic factor polymorphism Val66Met identifies fibromyalgia syndrome subgroup with higher body mass index and C-reactive protein.

A common single nucleotide polymorphism (SNP) in the gene of brain-derived neurotrophic factor (BDNF) results from a substitution at position 66 from valine (Val) to methionine (Met) and may predispose to human neuropsychiatric disorders. We proposed to determine whether these BDNF gene SNPs were associated with fibromyalgia syndrome (FMS) and/or any of its typical phenotypes. Patients with FMS (N = 95) and healthy normal controls (HNC, N = 58) were studied. Serum high-sensitivity C-reactive protein (hsCRP) levels were measured using an enzyme-linked immunosorbent assay (ELISA). The BDNF SNPs were determined using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP).The BDNF SNP distribution was 65 (68%) Val/Val, 28 (30%) Val/Met, and 2 (2%) Met/Met for FMS and 40 (69%), 17(29%), and 1 (2%) for HNC, respectively. The serum high-sensitivity C-reactive protein (hsCRP)and body mass index (BMI) in FMS were higher than in HNC. The FMS with BDNF Val66Val had significantly higher mean BMI (P = 0.0001) and hsCRP (P = 0.02) than did FMS carrying the Val66Met genotype. This pattern was not found in HNC. Phenotypic measures of subjective pain, pain threshold, depression, or insomnia did not relate to either of the BDNF SNPs in FMS. The relative distribution BDNF SNPs did not differ between FMS and HNC. The BDNF Val66Met polymorphism is not selective for FMS. The BDNF Val66Val SNP identifies a subgroup of FMS with elevated hsCRP and higher BMI. This is the first study to associate a BDNF polymorphism with a FMS subgroup phenotype.

Redox regulation of age-associated defects in generation and maintenance of T cell self-tolerance and immunity to foreign antigens.

Thymic atrophy reduces naive T cell production and contributes to increased susceptibility to viral infection with age. Expression of tissue-restricted antigen (TRA) genes also declines with age and has been thought to increase autoimmune disease susceptibility. We find that diminished expression of a model TRA gene in aged thymic stromal cells correlates with impaired clonal deletion of cognate T cells recognizing an autoantigen involved in atherosclerosis. Clonal deletion in the polyclonal thymocyte population is also perturbed. Distinct age-associated defects in the generation of antigen-specific T cells include a conspicuous decline in generation of T cells recognizing an immunodominant influenza epitope. Increased catalase activity delays thymic atrophy, and here, we show that it mitigates declining production of influenza-specific T cells and their frequency in lung after infection, but does not reverse declines in TRA expression or efficient negative selection. These results reveal important considerations for strategies to restore thymic function.

Age-Associated Decline in Thymic B Cell Expression of Aire and Aire-Dependent Self-Antigens.

Although autoimmune disorders are a significant source of morbidity and mortality in older individuals, the mechanisms governing age-associated increases in susceptibility remain incompletely understood. Central T cell tolerance is mediated through presentation of self-antigens by cells constituting the thymic microenvironment, including epithelial cells, dendritic cells, and B cells. Medullary thymic epithelial cells (mTECs) and B cells express distinct cohorts of self-antigens, including tissue-restricted self-antigens (TRAs), such that developing T cells are tolerized to antigens from peripheral tissues. We find that expression of the TRA transcriptional regulator Aire, as well as Aire-dependent genes, declines with age in thymic B cells in mice and humans and that cell-intrinsic and cell-extrinsic mechanisms contribute to the diminished capacity of peripheral B cells to express Aire within the thymus. Our findings indicate that aging may diminish the ability of thymic B cells to tolerize T cells, revealing a potential mechanistic link between aging and autoimmunity.

Normal cerebrospinal fluid levels of hypocretin-1 (orexin A) in patients with fibromyalgia syndrome.

BACKGROUND: The hypothalamic neuropeptide hypocretin (orexin) modulates sleep-wake, feeding and endocrine functions. Cerebrospinal fluid (CSF) hypocretin-1 (Hcrt-1) concentrations are low in patients with narcolepsy-cataplexy, a sleep disorder characterized by hypersomnolence and rapid eye movement (REM) sleep abnormalities. METHODS: We determined CSF Hcrt-1 concentrations of patients with the fibromyalgia syndrome (FMS), a condition characterized by fatigue, insomnia and in some cases daytime hypersomnolence. RESULTS: Basal CSF levels of Hcrt-1 in FMS did not differ from those in healthy normal controls. CONCLUSIONS: These findings suggest that abnormally low Hcrt-1 is not a likely cause of fatigue in FMS.

Buckling Reduces eNOS Production and Stimulates Extracellular Matrix Remodeling in Arteries in Organ Culture.

Artery buckling alters the fluid shear stress and wall stress in the artery but its temporal effect on vascular wall remodeling is poorly understood. The purpose of this study was to investigate the early effect of artery buckling on endothelial nitric oxide synthase (eNOS) expression and extracellular matrix remodeling. Bilateral porcine carotid arteries were maintained in an ex vivo organ culture system with and without buckling while under the same physiological pressure and flow rate for 3-7 days. Matrix metalloproteinase-2 (MMP-2), MMP-9, fibronectin, elastin, collagen I, III and IV, tissue inhibitor of metalloproteinase-2 (TIMP-2), and eNOS were determined using Western blotting and immunohistochemistry. Our results showed that MMP-2 expression level was significantly higher in buckled arteries than in the controls and higher at the inner curve than at the outer curve of buckled arteries, while collagen IV content showed an opposite trend, suggesting that artery buckling increased MMP-2 expression and collagen IV degradation in a site-specific fashion. However, no differences for MMP-9, fibronectin, elastin, collagen I, III, and TIMP-2 were observed among the outer and inner curve sides of buckled arteries and straight controls. Additionally, eNOS expression was significantly decreased in buckled arteries. These results suggest that artery buckling triggers uneven wall remodeling that could lead to development of tortuous arteries.

Methylation-induced silencing of miR-34a enhances chemoresistance by directly upregulating ATG4B-induced autophagy through AMPK/mTOR pathway in prostate cancer.

miR-34a is downregulated and a regulator of drug resistance in prostate cancer (PCa). However, the mechanism of miR-34a in chemoresistance of PCa remains largely unknown. In the present study, we first confirmed the hypermethylation­induced downregulation of miR-34a in PCa tissues and cell lines, PC-3 and DU145. Additionally, transfection of miR-34a mimics and demethylation by 5-azacytidine both resulted in the upregulation of miR-34a expression, which further induced declined cell proliferation and the enhanced apoptosis in PCa cells. Upregulation of miR-34a enhanced the chemosensitivity of PC-3 and DU145 cells. Furthermore, overexpression of miR-34a reduced the expression of autophagy-related proteins, ATG4B, Beclin-1 and LC3B II/I in PCa cells and demethylation treatment showed similar effect. ATG4B was confirmed directly by miR-34a targeting in PCa. Finally, downregulated p-AMPK and upregulated p-mTOR were detected in miR-34a overexpressed PCa cells. Collectively, miR-34a enhances chemosensitivity by directly downregulating ATG4B-induced autophagy through AMPK/mTOR pathway in PCa.

microRNA-32 induces radioresistance by targeting DAB2IP and regulating autophagy in prostate cancer cells.

The aberrant expression of microRNAs (miRNAs/miRs) has been found in numerous cancer types. miR-32 is an oncomiR in prostate cancer (PCa), however, the mechanisms by which miR-32 functions as a regulator of radiotherapy response and resistance in PCa are largely unknown. In the present study, it was found that DAB2 interacting protein (DAB2IP), the miR-32-dependent tumor-suppressor gene, was downregulated and induced autophagy and inhibited radiotherapy-induced apoptosis in PCa cells. miR-32 expression was upregulated or overexpressed in PCa, and miR-32 inhibited DAB2IP expression through a direct binding site within the DAB2IP 3' untranslated region. miR-32 mimics enhanced tumor cell survival and decreased radiosensitivity in the PCa cells, which were reversed by miR-32 inhibitor. Flow cytometric analysis revealed that overexpressed miR-32, consistent with the DAB2IP-knockdown results, reduced ionizing radiation (IR)-induced cell apoptosis, which was restored by 4 nM brefeldin A treatment. More significantly, the overexpression of miR-32 and the knockdown of DAB2IP enhanced autophagy in the IR-treated PCa cells. miR-32 regulated the expression of autophagy-related proteins, such as DAB2IP, Beclin 1 and Light chain 3ß I/II, as well as phosphorylation of S6 kinase and mammalian target of rapamycin. In conclusion, these data provide novel insights into the mechanisms governing the regulation of DAB2IP expression by miR-32 and their possible contribution to autophagy and radioresistance in PCa.

Artery Remodeling Under Axial Twist in Three Days Organ Culture.

Arteries often endure axial twist due to body movement and surgical procedures, but how arteries remodel under axial twist remains unclear. The objective of this study was to investigate early stage arterial wall remodeling under axial twist. Porcine carotid arteries were twisted axially and maintained for three days in ex vivo organ culture systems while the pressure and flow remained the same as untwisted controls. Cell proliferation, internal elastic lamina (IEL) fenestrae shape and size, endothelial cell (EC) morphology and orientation, as well as the expression of matrix metalloproteinases (MMPs), MMP-2 and MMP-9, and tissue inhibitor of metalloproteinase-2 (TIMP-2) were quantified using immunohistochemistry staining and immunoblotting. Our results demonstrated that cell proliferation in both the intima and media were significantly higher in the twisted arteries compared to the controls. The cell proliferation in the intima increased from 1.33 ± 0.21% to 7.63 ± 1.89%, and in the media from 1.93 ± 0.84% to 8.27 ± 2.92% (p < 0.05). IEL fenestrae total area decreased from 26.07 ± 2.13% to 14.74 ± 0.61% and average size decreased from 169.03 ± 18.85 µm(2) to 80.14 ± 1.96 µm(2) (p < 0.01), but aspect ratio increased in the twist group from 2.39 ± 0.15 to 2.83 ± 0.29 (p < 0.05). MMP-2 expression significantly increased (p < 0.05) while MMP-9 and TIMP-2 showed no significant difference in the twist group. The ECs in the twisted arteries were significantly elongated compared to the controls after three days. The angle between the major axis of the ECs and blood flow direction under twist was 7.46 ± 2.44 degrees after 3 days organ culture, a decrease from the initial 15.58 ± 1.29 degrees. These results demonstrate that axial twist can stimulate artery remodeling. These findings complement our understanding of arterial wall remodeling under mechanical stress resulting from pressure and flow variations.

Suppression of Cellular Proliferation and Invasion by HMGB1 Knockdown in Bladder Urothelial Carcinoma Cells.

HMGB1, which acts as a DNA chaperone to help maintain nuclear homeostasis, was reported to play a prominent role in cancer progression, angiogenesis, invasion, and metastasis development. Increased expression of HMGB1 has been observed in several tumor entities. However, the molecular mechanisms of HMGB1 in tumorigenesis of bladder cancer have rarely been reported. In the present study, real-time quantitative RT-PCR analysis revealed that the expression of HMGB1 in human bladder urothelial carcinoma (BUC) cells was much higher than that in human normal urethra epithelial cells. In order to investigate the role of HMGB1 in BUC cells, RNA interference and Talen-mediated gene knockout (KO) were used to knockdown and knockout HMGB1, respectively, in BUC cell lines BIU-87 and T24. HMGB1 knockdown/out greatly inhibited proliferation, invasion, and cell cycle G1/S transition of BUC cells. The decrease in cell viability caused by HMGB1 knockdown/out was due to an increase in apoptosis via Bax/Bcl-2, both of which were important molecules involved in the apoptotic pathway. We then investigated the effect of HMGB1 knockdown/out on the sensitivity of BUC cells treated with the anticancer drug cisplatin. Knockdown or knockout of HMGB1 rendered BUC cells more sensitive to cisplatin. The decreased expression of LC3-II and Beclin 1, which resulted in decreased levels of autophagy, could probably explain this phenomenon. Thus, HMGB1 may become a novel promising candidate for the prognosis and therapy for bladder cancer.

Mechanical instability of normal and aneurysmal arteries.

Tortuous arteries associated with aneurysms have been observed in aged patients with atherosclerosis and hypertension. However, the underlying mechanism is poorly understood. The objective of this study was to determine the effect of aneurysms on arterial buckling instability and the effect of buckling on aneurysm wall stress. We investigated the mechanical buckling and post-buckling behavior of normal and aneurysmal carotid arteries and aorta's using computational simulations and experimental measurements to elucidate the interrelationship between artery buckling and aneurysms. Buckling tests were done in porcine carotid arteries with small aneurysms created using elastase treatment. Parametric studies were done for model aneurysms with orthotropic nonlinear elastic walls using finite element simulations. Our results demonstrated that arteries buckled at a critical buckling pressure and the post-buckling deflection increased nonlinearly with increasing pressure. The presence of an aneurysm can reduce the critical buckling pressure of arteries, although the effect depends on the aneurysm's dimensions. Buckled aneurysms demonstrated a higher peak wall stress compared to unbuckled aneurysms under the same lumen pressure. We conclude that aneurysmal arteries are vulnerable to mechanical buckling and mechanical buckling could lead to high stresses in the aneurysm wall. Buckling could be a possible mechanism for the development of tortuous aneurysmal arteries such as in the Loeys-Dietz syndrome.

Smooth muscle cell contraction increases the critical buckling pressure of arteries.

Recent in vitro experiments demonstrated that arteries under increased internal pressure or decreased axial stretch may buckle into the tortuous pattern that is commonly observed in aging or diseased arteries in vivo. It suggests that buckling is a possible mechanism for the development of artery tortuosity. Vascular tone has significant effects on arterial mechanical properties but its effect on artery buckling is unknown. The objective of this study was to determine the effects of smooth muscle cell contraction on the critical buckling pressure of arteries. Porcine common carotid arteries were perfused in an ex vivo organ culture system overnight under physiological flow and pressure. The perfusion pressure was adjusted to determine the critical buckling pressure of these arteries at in vivo and reduced axial stretch ratios (1.5 and 1.3) at baseline and after smooth muscle contraction and relaxation stimulated by norepinephrine and sodium nitroprusside, respectively. Our results demonstrated that the critical buckling pressure was significantly higher when the smooth muscle was contracted compared with relaxed condition (97.3mmHg vs 72.9mmHg at axial stretch ratio of 1.3 and 93.7mmHg vs 58.6mmHg at 1.5, p<0.05). These results indicate that arterial smooth muscle cell contraction increased artery stability.

Effects of physical exercise on serum levels of serotonin and its metabolite in fibromyalgia: a randomized pilot study.

To evaluate the effects of aerobic training and stretching on serum levels of serotonin (5HT) and its main metabolite 5-hydroxindolacetic acid (5HIAA). Twenty-two women with FM were randomized into one of two exercise modalities (aerobic walking exercise or stretching exercise) to be accomplished three times a week for 20 weeks. The serum levels of 5HT and 5HIAA were evaluated before and after the exercise program by high performance liquid chromatography (HPLC) with colorimetric detection. Within group analysis (pre-post) showed that serum levels of both 5HT and 5HIAA changed significantly in the aerobic group during the 20-week course of therapy (5HT: P = 0,03; 5HIAA: P = 0,003). In the stretching group, however, no statistically significant change was observed (5HT: P=0,491; 5HIAA: P=0,549). Between group statistical comparisons of laboratory measures disclosed that aerobic training was superior to stretching in that it significantly increased the levels of 5HIAA (F test = 6.61; P = 0.01), but the average difference between groups on the levels of 5HT did not meet significance criteria (F test = 3.42; P = 0.08). Aerobic training increases the 5HIAA and 5HT levels and it could explain why aerobic exercise can improve symptoms in fibromyalgia syndrome patient more than stretching exercise.

Alterations in Pulse Pressure Affect Artery Function.

Pulse pressure changes in response to cardiovascular diseases and interventions, but its effect on vascular wall structure and function is poorly understood. We examined the effect of increased or decreased pulse pressure on artery function, cellular function, and extracellular matrix remodeling. Porcine carotid arteries were cultured under non-pulsatile (100 mmHg), pulsatile (70-130 mmHg), or hyper-pulsatile pressure (50-150 mmHg) for 1 to 3 days. Vasomotor response, wall permeability, cell proliferation, apoptosis, extracellular matrix remodeling, and proteins involved in atherogenesis were examined. Our results showed that hyper-pulsatile pressure decreased the artery response to sodium nitroprusside, basal tone, and wall permeability after three days. Non-pulsatile pressure increased cell proliferation. Neither hyper-pulsatile nor non-pulsatile pressure caused a change in the extracellular matrix or in the expression of matrix metalloproteinase-2 (MMP-2), MMP-9, caveolin-1, or α-actin. Hyper-pulsatile pressure increased monocyte chemotactic protein-1 gene expression. Taken together, these changes indicate that pulse pressure has a limited effect on the artery immediately after its application. Specifically an increase in pulse pressure alters the artery tone and wall permeability while a decrease in pulse pressure alters cell proliferation. Overall these results provide insight into how the artery initially responds to changes in pulse pressure.

Genetic polymorphisms of the beta2-adrenergic receptor relate to guanosine protein-coupled stimulator receptor dysfunction in fibromyalgia syndrome.

OBJECTIVE: To determine the genotype frequencies of ß(2)-adrenergic receptor (ß(2)AR) gene polymorphisms (Gly16Arg, Glu27Gln) in patients with fibromyalgia syndrome (FM) by comparison with unrelated healthy controls. We sought any clinical association with these polymorphisms and determined whether the polymorphisms would associate with a biologic guanosine protein-coupled stimulator receptor (Gs) dysfunction in FM. METHODS: Study subjects included 97 clinically characterized patients with FM and 59 controls. The ß(2)AR polymorphisms at codons 16 and 27 were determined using polymerase chain reaction-restriction fragment length polymorphism. The Gs functions of peripheral blood mononuclear cells (PBMC) were tested using isoproterenol (ISO) as the adrenergic Gs ligand and measuring intracellular cyclic adenosine monophosphate (cAMP) levels. RESULTS: The frequency of the ß(2)AR gene polymorphism Gly16Arg in FM (43.5%) was significantly lower than in controls (63.2%), suggesting that this genotype might have some effect on the risk of developing FM. The only clinical association in FM was with sleep dysfunction. Patients with FM who carried the ß(2)AR polymorphism Arg16Arg also exhibited significantly lower PBMC basal cAMP levels (p < 0.05) and lower ISO-stimulated cAMP levels (p < 0.05) than FM carrying Gly16Gly or Gly16Arg. CONCLUSION: This confirms a relationship between ß(2)AR polymorphism and FM. It is the first study to demonstrate ß(2)AR polymorphism-related differences in intracellular cAMP responses of FM PBMC after ß(2)AR stimulation in vitro. These findings may explain some of the differences in responsiveness of FM subgroups to the adrenergic agonist medications currently approved for FM treatment.

NF-kappa B activation is not required for Chlamydia trachomatis inhibition of host epithelial cell apoptosis.

Chlamydia trachomatis, an obligate intracellular bacterial species, is known to inhibit host cell apoptosis. However, the chlamydial antiapoptotic mechanism is still not clear. Because NF-kappaB activation is antiapoptotic, we tested the potential role of NF-kappaB activation in chlamydial antiapoptotic activity in the current study. First, no obvious NF-kappaB activation was detected in the chlamydia-infected cells when these cells were resistant to apoptosis induced via either the intrinsic or extrinsic apoptosis pathways. Second, inhibition of NF-kappaB activation with pharmacologic reagents failed to block the chlamydial antiapoptotic activity. Finally, NF-kappaB p65 gene deletion did not prevent chlamydia from inhibiting host cell apoptosis. These observations together have demonstrated that NF-kappaB activation is not required for the chlamydial antiapoptotic activity.

Degradation of the proapoptotic proteins Bik, Puma, and Bim with Bcl-2 domain 3 homology in Chlamydia trachomatis-infected cells.

We have previously correlated Chlamydia trachomatis antiapoptotic activity with the blockade of mitochondrial cytochrome c release and the inhibition of Bax and Bak activation. We now report that C. trachomatis infection leads to degradation of Bik, Puma, and Bim, three upstream proapoptotic BH3-only proteins of the Bcl-2 family that can transmit death signals to mitochondria by inhibiting the Bcl-2 antiapoptotic proteins and/or activating the Bcl-2 proapoptotic members, such as Bax and Bak. This observation has provided new information on the chlamydial antiapoptosis mechanisms.