Endothelin-1 causes a prolonged protein kinase C activation and acts as a co-mitogen in vascular smooth muscle cells

Endothelin-1 (ET-1) is known to act via G-protein coupled receptors. It has therefore been suggested that any mitogenic activity it may possess, is due to activation of phospholipase C and protein kinase C (PKC). We have therefore examined both the ability of ET-1 to act as a mitogen and its ability to activate PKC. We found that ET-1 significantly increased thymidine incorporation and enhanced platelet-derived growth factor-induced DNA synthesis, as well as causing a prolonged translocation of PKC to the cell membrane. ET-1 significantly increased PKC dependent phosphorylation of two specific substrates. The phosphorylation of MBP_4–14 (from myelin basic protein) was partially dependent on extracellular Ca²⁺, implicating activation of PKC-α, whereas phosphorylation of the so called ε-peptide was Ca²⁺-independent and prolonged. This could be due either to the δ or ζ isoform of PKC, known to be present in these cells. However, ET-1 induced little proliferation or PKC activity in a transformed smooth muscle cell line, DDT₁ MF-2, which lacks expression of the PKC-αisoform, but expresses the ζ-isoform. Thus, it would appear that ET-1-induced mitogenicity in smooth muscle cells may be related to the sustained, Ca²⁺-independent activation of PKC-δ.     

Adhesion dependent release of hepatocyte growth factor and interleukin-1 receptor antagonist from human blood granulocytes and monocytes: Evidence for the involvement of plasma IgG,complement C3 and β2 integrin

Objective:Evolving evidence of anti-inflammatory effects is observed in patients with rheumatoid arthritis or ulcerative colitis following periodic adsorptive granulocyte and monocyte (GM) apheresis with a column containing cellulose acetate (CA) beads as apheresis carriers. This study was undertaken to obtain insights into mechanisms of anti-inflammatory actions of adsorptive GM apheresis with CA beads. Methods:In a series of in-vitro experiments, we investigated the effects of plasma proteins and the leucocytes 2 integrin (CD18) on granulocyte adsorption to CA beads. Results:Granulocyte adsorption to CA beads required plasma IgG, the complement C3 and was inhibited by an antibody to leucocytes CD18. Further, hepatocyte growth factor (HGF) and interleukin-1 receptor antagonist (IL-1ra) which have strong anti-inflammatory actions were released by granulocytes that adhered to CA beads. Conclusions:Plasma IgG, C3 derived complement activation fragments and leucocytes CD18 are involved in granulocyte adhesion to CA beads and hence the release of HGF and IL-1ra.Received 27 October 2003; returned for revision 16 December 2003; accepted by M. J. Parnham 8 January 2004     

Regulation of membrane excitability by intracellular pH (pHi) changers through Ca2+-activated K+ current (BK channel) in single smooth muscle cells from rabbit basilar artery

Employing microfluorometric system and patch clamp technique in rabbit basilar arterial myocytes, regulation mechanisms of vascular excitability were investigated by applying intracellular pH (pH_i) changers such as sodium acetate (SA) and NH₄Cl. Applications of caffeine produced transient phasic contractions in a reversible manner. These caffeine-induced contractions were significantly enhanced by SA and suppressed by NH₄Cl. Intracellular Ca²⁺ concentration ([Ca²⁺]_i) was monitored in a single isolated myocyte and based the ratio of fluorescence using Fura-2 AM (R _340/380). SA (20 mM) increased and NH₄Cl (20 mM) decreased R _340/380 by 0.2 ± 0.03 and 0.1 ± 0.02, respectively, in a reversible manner. Caffeine (10 mM) transiently increased R _340/380 by 0.9 ± 0.07, and the ratio increment was significantly enhanced by SA and suppressed by NH₄Cl, implying that SA and NH₄Cl may affect [Ca²⁺]_i (p < 0.05). Accordingly, we studied the effects of SA and NH₄Cl on Ca²⁺-activated K⁺ current (IK_Ca) under patch clamp technique. Caffeine produced transient outward current at holding potential (V _h) of 0 mV, caffeine induced transient outward K⁺ current, and the spontaneous transient outward currents were significantly enhanced by SA and suppressed by NH₄Cl. In addition, IK_Ca was significantly increased by acidotic condition when pH_i was lowered by altering the NH₄Cl gradient across the cell membrane. Finally, the effects of SA and NH₄Cl on the membrane excitability and basal tension were studied: Under current clamp mode, resting membrane potential (RMP) was −28 ± 2.3 mV in a single cell level and was depolarized by 13 ± 2.4 mV with 2 mM tetraethylammonium (TEA). SA hyperpolarized and NH₄Cl depolarized RMP by 10 ± 1.9 and 16 ± 4.7 mV, respectively. SA-induced hyperpolarization and relaxation of basal tension was significantly inhibited by TEA. These results suggest that SA and NH₄Cl might regulate vascular tone by altering membrane excitability through modulation of [Ca²⁺]_i and Ca²⁺-activated K channels in rabbit basilar artery.     

Nicotine modifies in vivo and in vitro rat hippocampal amyloid precursor protein processing in young but not old rats

Previous studies have shown that administration of nicotine modifies the expression and secretion of amyloid precursor protein (APP) in various cell lines. The present study investigated the extent to which chronic subcutaneous nicotine administration influences APP levels and processing in cerebral cortex, striatum and hippocampus of young and old rat brains. The results showed that constant nicotine infusion (0.25 or 4.00 mg/kg/day) increased the levels of particulate APP (APPp) but not secreted APP (APPs) in the hippocampus of young rats in vivo. This response to nicotine was not observed in the striatum or cerebral cortex of young rats or in any of the brain regions examined in old animals. Subsequent in vitro analysis demonstrated that nicotine enhanced the release of APPs from hippocampal slice preparations and that this increase was attenuated by mecamylamine, a non-selective nicotinic acetylcholine receptor (nAChR) antagonist. The in vitro effect of nicotine on APPs was age-related, being only detected from hippocampal slices derived from the young but not the older animals. These results suggest that nicotine modulates APP expression and secretion in the hippocampus and that the responses observed to the drug are age-dependent being only detected in younger rats.     

Upregulation of HTLV-1 and HTLV-2 expression by HIV-1 in vitro

Co-infections with HIV-1 and the human T leukemia virus types 1 and 2 (HTLV-1, HTLV-2) occur frequently, particularly in large metropolitan areas where injection drug use is a shared mode of transmission. Recent evidence suggests that HIV-HTLV co-infections are associated with upregulated HTLV-1/2 virus expression and disease. An in vitro model of HIV-1 and HTLV-1/2 co-infection was utilized to determine if cell free HIV-1 virions or recombinant HIV-1 Tat protein (200-1,000 ng/ml) upregulated HTLV-1/2 expression and infectivity. Exposure to HIV-1 increased the number of HTLV-1 antigen expressing cells, from 6% at baseline to 12% at 24 hr, and 20% at 120 hr (P < 0.05) post-exposure. A similar, although less robust response was observed in HTLV-2 infected cells. HIV-1 co-localized almost exclusively with HTLV-1/2 positive cells. Exposure to HIV-1 Tat protein (1,000 ng/ml) increased HTLV-1 p19 expression almost twofold by 48 hr, and cells co-stimulated with 10 nM phorbol myristate acetate (PMA) showed almost a fourfold increase over baseline. It is concluded that HIV-1 augments HTLV-1/2 infectivity in vitro. The findings also suggest a role for the HIV-1 Tat protein and PMA-inducible cellular factors, in HIV-1 induced HTLV-1/2 antigen expression.     

Immediate effects of foot orthoses on pain during functional tasks in people with patellofemoral osteoarthritis: A cross-over,proof-of-concept study

Background

The purpose of the study was to determine whether prefabricated foot orthoses immediately reduce pain during functional tasks in people with patellofemoral osteoarthritis, compared to flat insoles and shoes alone.

Prevention of unintended pregnancies by using emergency contraception: the differences between levonorgestrel and ulipristal acetate. A theoretical model using data from a survey on the use of emergency contraception in Spain, 2017

Objective: To estimate the differences in unintended pregnancies avoided using either levonorgestrel (LNG) or ulipristal acetate (UPA) emergency contraception (EC).

Design: Cross-sectional study.

Setting: Survey carried out in Spain.

Participants: 1000 Spanish women reporting unprotected sex in 2017.

Main measurements: EC use, reasons for not using EC, calculation of the number of unintended pregnancies avoided.

Results: 39% of Spanish women having had unprotected sex used EC. 61% of those women did not use EC and 11% did not know the existence of this resource. In 2017 the use of EC prevented 101,271 unintended pregnancies. If instead of using LNG every woman had used UPA another 15,979 additional pregnancies could have been prevented.

Conclusions: If all Spanish women having unprotected sex used EC we could expect a significant decrease in the number of unintended pregnancies and abortions. Using UPA instead of LNG would have a greater impact on that reduction with the corresponding benefit for women and society as a whole.     

Decreased tissue inhibitor of metalloproteinase in the endometrium of women using depot medroxyprogesterone acetate: a role for altered endometrial matrix metalloproteinase/tissue inhibitor of metalloproteinase balance in the pathogenesis of abnormal uterine bleeding?

BACKGROUND: Abnormal uterine bleeding is commonly associated with progestin-only contraceptives, including depot medroxyprogesterone acetate (DMPA), and remains the main reason why these agents are discontinued. Matrix metalloproteinases (MMP), enzymes which degrade specific extracellular matrix components, and leukocytes are implicated in menstruation. Alteration in endometrial MMP-9 and leukocytes has been described in users of other progestin-only contraceptives, suggesting a potential role in the pathogenesis of abnormal uterine bleeding. METHODS: This study describes the immunohistochemical localization of MMP-9, the tissue inhibitors of metalloproteinases (TIMP)-1, TIMP-2 and TIMP-3, and leukocytes [CD3+ T lymphocytes, CD68+ macrophages and CD56+ uterine natural killer cells (uNK cells)] in the endometrium of women using DMPA. Comparison is made with perimenstrual endometria from normal cycling women. RESULTS: Similar to the perimenstrual period, an influx of MMP-9 positive cells (identified as neutrophils and CD3+ T cells on the basis of dual immunofluorescence), macrophages and uNK cells was observed in the endometrium of DMPA users. However, significantly more endometrial T lymphocytes were observed in DMPA users. Immunoreactive TIMP, present in all endometrial compartments, demonstrated a significantly decreased immunostaining intensity score in endometrial epithelium (TIMP-1 and TIMP-2), stroma (TIMP-1, TIMP-2 and TIMP-3), endothelium (TIMP-1 and TIMP-2) and vascular smooth muscle (TIMP-1) of DMPA users compared with controls. No correlation was observed between the parameters studied and bleeding patterns reported by subjects. CONCLUSIONS: These findings provide additional evidence for the importance of the MMP/TIMP balance in the loss/maintenance of endometrial integrity and in the complex pathological mechanisms involved in the troubling side-effect of menstrual bleeding disturbance.     

Chain‐Length‐Dependent Termination of Vinyl Acetate and Vinyl Pivalate Bulk Homopolymerizations Studied by SP‐PLP‐EPR

Bulk homopolymerizations of vinyl acetate and vinyl pivalate are studied by EPR experiments between ?65 °C and 60 °C with dicumyl peroxide acting as the photoinitiator. No mid‐chain radicals are seen, which demonstrates that backbiting plays no role. The chain‐length dependence of the termination rate coefficients measured up to 13% monomer conversion is adequately represented by the composite model. The power‐law exponents α_s and α_l for short‐chain and long‐chain radicals are: α_s(VAc) = 0.57 ± 0.05, α_s(VPi) = 0.67 ± 0.15, α_l(VAc) = 0.16 ± 0.07, and α_l(VPi) = 0.16 ± 0.07. The crossover chain lengths differ largely: i_c(VAc) = 20 ± 10 and i_c(VPi) = 110 ± 30. The rate coefficient for termination of two radicals of chain length unity, , which is the fourth composite‐model parameter, depends on temperature, as does the monomer fluidity.