RAS induced senescence of skin keratinocytes is mediated through Rho-associated protein kinase (ROCK).

Cellular senescence is a well-documented response to oncogene activation in many tissues. Multiple pathways are invoked to achieve senescence indicating its importance to counteract the transforming activities of oncogenic stimulation. We now report that the Rho-associated protein kinase (ROCK) signaling pathway is a critical regulator of oncogene-induced senescence in skin carcinogenesis. Transformation of mouse keratinocytes with oncogenic RAS upregulates ROCK activity and initiates a senescence response characterized by cell enlargement, growth inhibition, upregulation of senescence associated ß-galactosidase (SAßgal) expression, and release of multiple pro-inflammatory factors comprising the senescence-associated secretory phenotype (SASP). The addition of the ROCK inhibitor Y-27632 and others prevents these senescence responses and maintains proliferating confluent RAS transformed keratinocyte cultures indefinitely. Mechanistically, oncogenic RAS transformation is associated with upregulation of cell cycle inhibitors p15Ink4b , p16Ink4a , and p19Arf and downregulation of p-AKT, all of which are reversed by Y-27632. RNA-seq analysis of Y-27632 treated RAS-transformed keratinocytes indicated that the inhibitor reduced growth-inhibitory gene expression profiles and maintained expression of proliferative pathways. Y-27632 also reduced the expression of NF-κB effector genes and the expression of IκBζ downstream mediators. The senescence inhibition from Y-27632 was reversible, and upon its removal, senescence reoccurred in vitro with rapid upregulation of cell cycle inhibitors, SASP expression, and cell detachment. Y-27632 treated cultured RAS-keratinocytes formed tumors in the absence of the inhibitor when placed in skin orthografts suggesting that factors in the tumor microenvironment can overcome the drive to senescence imparted by overactive ROCK activity.

Botulinum toxin A (Botox) injection into muscles of pelvic floor as a treatment for persistent pelvic pain secondary to pelvic floor muscular spasm: A pilot study.

BACKGROUND: Persistent pelvic pain (PPP) remains an important cause of morbidity. Pelvic floor muscle spasm is an important contributor to PPP. AIMS: The study's primary aim was to assess if botulinum toxin (BoNT) injection to pelvic floor muscles altered pain scores or quality of life (QoL) at six, 12 and 26 weeks. Secondary aims included investigating the impact of BoNT on opiate usage, examining the role of pain catastrophising, and assessing for complications. MATERIALS AND METHODS: A single-centre prospective cohort study enrolled 21 patients with PPP who had failed physiotherapy techniques. Each participant underwent BoNT injection to muscles of the pelvic floor and pudendal nerve block. Questionnaires and digital vaginal examinations were conducted at baseline, six, 12 and 26 weeks. Pain score quantification used visual analogue scales (VAS) and numerical rating scales (NRS). Other outcome assessments included The World Health Organization Quality of Life instrument (WHOQoL-BREF), Pain Catastrophising Scale (PCS), and modified Australian Pelvic Floor Questionnaire (APFQ). ACTRN12620000067976. RESULTS: Following BoNT injection, median VAS scores decreased for all domains at six and 12 weeks, with VAS for dyspareunia significant at six weeks (P = 0.026). Scores returned to baseline by 26 weeks. Opiate usage was significantly less following BoNT injection, with a percentage reduction of 23.8% (95% CI -48.3 to 0.7, P = 0.06). Sexual function improved significantly (P < 0.01), and at six months, four previously apareunic participants reported successful penetrative vaginal intercourse. Health-related QoL and PCS demonstrated sustained improvement (P = 0.02-0.05). NRS for muscle tenderness decreased for all assessed muscle groups (P < 0.001). CONCLUSIONS: BoNT requires further assessment as a treatment modality for select women with PPP.

Topical Application of a Dual ABC Transporter Substrate and NF-κB Inhibitor Blocks Multiple Sources of Cutaneous Inflammation in Mouse Skin.

Among the molecular signals underlying cutaneous inflammation is the transcription complex NF-κB, its upstream modulators, and cytokines and chemokines that are the downstream proinflammatory effectors. Central to NF-κB activation is IκB kinase (IKK), which phosphorylates IκBα, releasing NF-κB to the nucleus. In a screening of a kinase inhibitor library, we identified two IKK inhibitors that were high-affinity substrates for p-glycoprotein (ABCB1), the multidrug resistance protein known to facilitate transdermal drug delivery. ACHP (2-amino-6-[2-(cyclopropylmethoxy)-6-hydroxyphenyl]-4-(4-piperidinyl)-3-pyridinecarbonitrile) and IKK 16 prevented both nuclear translocation of NF-κB and activation of a NF-κB reporter and reduced the induction of cytokine and chemokine transcripts in human or mouse keratinocytes by IL-1α, tumor necrosis factor-α, and phorbol myristate acetate. ACHP, but not IKK 16, was nontoxic to mouse or human keratinocytes at any dose tested. In mice, topical ACHP prevented the cutaneous inflammation induced by topical phorbol myristate acetate or imiquimod, reduced the inflammation from erythema doses of artificial sunlight, and lowered the tumor incidence of mice treated with 7,12-dimethyl benzanthracene when applied before phorbol myristate acetate. Topical ACHP also reduced the NF-κB and IL-17 inflammatory signature after multiple doses of imiquimod. Thus, ACHP and IKK 16 hit their NF-κB target in mouse and human keratinocytes, and ACHP is an effective topical nonsteroidal anti-inflammatory in mice.

Regional and Facility Differences in Interventions for Mastitis by Australian Physiotherapists.

BACKGROUND: Little information has been documented regarding interventions for mastitis by Australian physiotherapists. It is currently not known if physiotherapy interventions vary across Australian regions and types of healthcare facilities. RESEARCH AIMS: (1) To identify the interventions used by Australian physiotherapists treating mothers with mastitis and (2) to determine the variability in interventions used across regions and facilities. METHODS: A retrospective observational design was used. A sample of case records of mothers with mastitis was identified (N = 192). These case records documented physiotherapy interventions for mastitis in hospitals and private physiotherapy practices in Western Australia (n = 77; 40.1%), Victoria (n = 76; 39.6%), and New South Wales (n = 39; 20.3%). An electronic data collection tool was designed to examine intervention variables. RESULTS: The physiotherapy interventions received by mothers included therapeutic ultrasound (n=175; 91.1%), education and advice (n = 160; 83.3%), and massage (n = 103; 53.6%). Therapeutic ultrasound parameters varied across regions and types of healthcare facilities. Mean documented therapeutic ultrasound intensity was approximately twice as high in New South Wales and Victoria than in Western Australia. CONCLUSIONS: Regional and facility differences exist in physiotherapy interventions for mastitis in Australia. Healthcare professionals who refer to physiotherapists for mastitis should be aware that interventions received may differ across regions and facility types.

Testing the feasibility of a mobile technology intervention promoting healthy gestational weight gain in pregnant women (txt4two) - study protocol for a randomised controlled trial.

BACKGROUND: Overweight, obesity and excess gestational weight gain (GWG) are associated with negative health outcomes for mother and child in pregnancy and across the life course. Interventions promoting GWG within guidelines report mixed results. Most are time and cost intensive, which limits scalability. Mobile technologies (mHealth) offer low cost, ready access and individually-tailored support. We aim to test the feasibility of an mHealth intervention promoting healthy nutrition, physical activity and GWG in women who begin pregnancy overweight or obese. METHODS/DESIGN: txt4two is a parallel randomised control trial pilot recruiting women with a singleton, live gestation between 10(+0) and 17(+6) weeks at the first hospital antenatal clinic visit. Inclusion criteria are pre-pregnancy BMI > 25 kg/m(2) and mobile phone ownership. One hundred consenting women will be randomised to intervention or control groups at a 1:1 ratio. All participants will receive standard antenatal care. In addition, the txt4two intervention will be delivered from baseline to 36 weeks gestation and consists of a tailored suite of theoretically-grounded, evidence-based intervention strategies focusing on healthy nutrition, physical activity and GWG. This includes: mobile phone interactive text messages promoting positive health behaviours, goal setting and self-monitoring; video messages; an information website; and a private moderated Facebook® chat forum. The primary outcome is the feasibility of the intervention. Secondary outcomes include GWG and participants' knowledge and behaviour regarding diet and physical activity during pregnancy. DISCUSSION: Findings will inform the development of larger-scale mHealth programmes to improve the delivery of healthy pregnancy nutrition, physical activity and GWG, that could be widely translated and disseminated. TRIAL REGISTRATION: Australian New Zealand Clinical Trials Registry: ACTRNU111111544397 . Date of registration: 19 March 2014.

Overexpression of miR­145 in U87 cells reduces glioma cell malignant phenotype and promotes survival after in vivo implantation.

In the present study, we sought to elucidate the effect of miR­145 on glioma cell progression and its mechanisms of action. We examined the effects of miR­145 on proliferation and invasion of U87 glioma cells and on capillary tube formation. Our data show that restoration of miR­145 in U87 glioma cells significantly reduced their in vitro proliferation, invasion and angiogenesis. However, decreased miR­145 expression promoted U87 glioma cell proliferation, invasion and angiogenesis, and reduced-expression of miR­145 increased ADAM17 and EGFR expression in U87 cells. Overexpression of miR­145 reduced ADAM17 and EGFR expression. VEGF secretion and VEGF expression were decreased by increased miR­145 expression in U87 cells and were reversed by miR­145 downregulation in vitro. Nude mice with intracerebral implantation of U87 overexpressing miR­145 cells exhibited significantly reduced tumor growth and promoted survival compared with control groups. Taken together, these results suggest a role for miR­145 as a tumor suppressor which inhibits glioma cell proliferation, invasion and angiogenesis in vitro and reduces glioma growth in vivo.