BNIP3 supports melanoma cell migration and vasculogenic mimicry by orchestrating the actin cytoskeleton

BNIP3 is an atypical BH3-only member of the BCL-2 family of proteins with reported pro-death as well as pro-autophagic and cytoprotective functions, depending on the type of stress and cellular context. In line with this, the role of BNIP3 in cancer is highly controversial and increased BNIP3 levels in cancer patients have been linked with both good as well as poor prognosis. In this study, using small hairpin RNA (shRNA) lentiviral transduction to stably knockdown BNIP3 (BNIP3-shRNA) expression levels in melanoma cells, we show that BNIP3 supports cancer cell survival and long-term clonogenic growth. Although BNIP3-shRNA increased mitochondrial mass and baseline levels of reactive oxygen species production, which are features associated with aggressive cancer cell behavior, it also prevented cell migration and completely abolished the ability to form a tubular-like network on matrigel, a hallmark of vasculogenic mimicry (VM). We found that this attenuated aggressive behavior of these melanoma cells was underscored by severe changes in cell morphology and remodeling of the actin cytoskeleton associated with loss of BNIP3. Indeed, BNIP3-silenced melanoma cells displayed enhanced formation of actin stress fibers and membrane ruffles, while lamellopodial protrusions and filopodia, tight junctions and adherens junctions were reduced. Moreover, loss of BNIP3 resulted in re-organization of focal adhesion sites associated with increased levels of phosphorylated focal adhesion kinase. Remarkably, BNIP3 silencing led to a drop of the protein levels of the integrin-associated protein CD47 and its downstream signaling effectors Rac1 and Cdc42. These observations underscore that BNIP3 is required to maintain steady-state levels of intracellular complexes orchestrating the plasticity of the actin cytoskeleton, which is integral to cell migration and other vital processes stimulating cancer progression. All together these results unveil an unprecedented pro-tumorigenic role of BNIP3 driving melanoma cell's aggressive features, like migration and VM

Comparison of phenotypic and genotypic tropism determination in triple-class-experienced HIV patients eligible for maraviroc treatment

BACKGROUND: Determination of HIV-1 tropism is a pre-requisite to the use of CCR5 antagonists. This study evaluated the potential of population genotypic tropism tests (GTTs) in clinical practice, and the correlation with phenotypic tropism tests (PTTs) in patients accessing routine HIV care. METHODS: Forty-nine consecutive plasma samples for which an original Trofile(TM) assay was performed were obtained from triple-class-experienced patients in need of a therapy change. Viral tropism was defined as the consensus of three or more tropism calls obtained from the combination of two independent population PTT assays (Trofile Biosciences, San Francisco, CA, USA, and Virco, Beerse, Belgium), population GTTs and GTTs based on ultra-deep sequencing. If no consensus was reached, a clonal PTT was performed in order to finalize the tropism call. This two-step approach allowed the definition of a reference tropism call. RESULTS: According to the reference tropism result, 35/49 samples were CCR5 tropic (R5) (patients eligible for maraviroc treatment) and 14/49 were assigned as non-R5 tropic. The non-R5 samples [patients not eligible for maraviroc treatment according to the FDA/European Medicines Agency (EMEA) label] group included both the CXCR4 (X4) samples and the dual and mixed CCR5/CXCR4 (R5/X4) samples. Compared with Trofile(TM) population PTTs, population GTTs showed a higher sensitivity (97%) and a higher negative predictive value (91%), but almost equal specificity and an equal positive predictive value. CONCLUSIONS: In line with recent reports from clinical trial data, our data support the use of population genotypic tropism testing as a tool for tropism determination before the start of maraviroc

Symphysiotomy in Zimbabwe; Postoperative Outcome, Width of the Symphysis Joint, and Knowledge, Attitudes and Practice among Doctors and Midwives

BACKGROUND: Obstructed labour remains one of the leading causes of maternal and foetal death and morbidity in poorly resourced areas of the world, where the 24 hours availability of a caesarean section (CS) cannot be guaranteed, and the CS related mortality rate is still high. In these settings, reinstatement of symphysiotomy has been advocated. The objectives were, in1994; to study perioperative and long-term complications of symphysiotomy and compare them to those related to CS for similar indications, in 1996; to measure the symphyseal width after symphysiotomy and compare it to that after normal vaginal delivery, and, in 1998; to assess knowledge, attitudes and practice related to symphysiotomy among doctors and midwives in Zimbabwe. METHODS AND FINDINGS: Thirty-four women who had undergone symphysiotomy and 29 women who had undergone a CS for obstructed labour were interviewed. The symphyseal widths of 19 women with a previous symphysiotomy were compared to that of 92 women with previous normal vaginal deliveries, using ultrasound technique. Forty-one doctors and 39 midwives, in three central hospitals and seven district hospitals in Zimbabwe, were interviewed about symphysiotomy. None of the 34 women reported serious soft tissue injuries or infections post symphysiotomy. Long-term complications after symphysiotomy do not differ notably from those after CS for similar indications. The intra-articular width of the symphysis pubis is increased after a symphysiotomy. Seventy-nine of the 80 interviewed health care workers knew about symphysiotomy. One obstetrician had performed symphysiotomies. Two-thirds of the participants considered symphysiotomy an obsolete and second-class operation, but lifesaving and appropriate in remote areas of Zimbabwe. Ten of 13 midwives in remote areas wanted to carry out symphysiotomies themselves. CONCLUSIONS: No severe complications due to symphysiotomy were revealed in this study. The results suggest that a modest permanent enlargement of the pelvis post symphysiotomy (together with the absence of a scarred uterus) may facilitate subsequent vaginal delivery. Doctors and midwives working in district hospitals have a more positive attitude to symphysiotomies than the colleagues in central hospitals. Obstetricians (who would have to do the teaching), working in the large urban hospitals almost exclude symphysiotomy as an alternative management in Zimbabwe

Implementation of a program for type 2 diabetes based on the Chronic Care Model in a hospital-centered health care system: "the Belgian experience"

Background: Most research publications on Chronic Care Model (CCM) implementation originate from organizations or countries with a well-structured primary health care system. Information about efforts made in countries with a less well-organized primary health care system is scarce. In 2003, the Belgian National Institute for Health and Disability Insurance commissioned a pilot study to explore how care for type 2 diabetes patients could be organized in a more efficient way in the Belgian healthcare setting, a setting where the organisational framework for chronic care is mainly hospital-centered. Methods: Process evaluation of an action research project (2003-2007) guided by the CCM in a well-defined geographical area with 76,826 inhabitants and an estimated number of 2,300 type 2 diabetes patients. In consultation with the region a program for type 2 diabetes patients was developed. The degree of implementation of the CCM in the region was assessed using the Assessment of Chronic Illness Care survey (ACIC). A multimethod approach was used to evaluate the implementation process. The resulting data were triangulated in order to identify the main facilitators and barriers encountered during the implementation process. Results: The overall ACIC score improved from 1.45 (limited support) at the start of the study to 5.5 (basic support) at the end of the study. The establishment of a local steering group and the appointment of a program manager were crucial steps in strengthening primary care. The willingness of a group of well-trained and motivated care providers to invest in quality improvement was an important facilitator. Important barriers were the complexity of the intervention, the lack of quality data, inadequate information technology support, the lack of commitment procedures and the uncertainty about sustainable funding. Conclusion: Guided by the CCM, this study highlights the opportunities and the bottlenecks for adapting chronic care delivery in a primary care system with limited structure. The study succeeded in achieving a considerable improvement of the overall support for diabetes patients but further improvement requires a shift towards system thinking among policy makers. Currently primary care providers lack the opportunities to take up full responsibility for chronic care

Application of AFLP for taxonomic and epidemiological studies of <i>Photobacterium damselae</i> subsp. <i>piscicida</i>

A collection of 106 Photobacterium damselae subsp. piscicida strains and 19 Photobacterium damselae subsp. damselae strains, including reference and type strains, were genetically characterized using AFLP. The total genomic DNA of each bacterial strain was digested using restriction endonucleases HindIII and TaqI. Using numerical analysis, six clusters were recognized. The largest cluster (n=106) contained the majority of the strains tested and consisted exclusively of Photobacterium damselae subsp. piscicida. The Photobacterium damselae subsp. damselae strains fell outside this cluster. DNA-DNA hybridization experiments showed 77% DNA binding between the two subspecies, indicating a close genetic relationship. This clearly demonstrates the applicability of AFLP in studying the taxonomic position of Photobacterium damselae subsp. piscicida. In addition, AFLP proved to be a useful genotypic technique for epidemiological surveys of the pathogen, since it was able to discriminate between Mediterranean and Japanese Photobacterium damselae subsp. piscicida isolates