Breast Tumor Metastasis and Its Microenvironment: It Takes Both Seed and Soil to Grow a Tumor and Target It for Treatment.

Metastasis remains a major challenge in treating breast cancer. Breast tumors metastasize to organ-specific locations such as the brain, lungs, and bone, but why some organs are favored over others remains unclear. Breast tumors also show heterogeneity, plasticity, and distinct microenvironments. This contributes to treatment failure and relapse. The interaction of breast cancer cells with their metastatic microenvironment has led to the concept that primary breast cancer cells act as seeds, whereas the metastatic tissue microenvironment (TME) is the soil. Improving our understanding of this interaction could lead to better treatment strategies for metastatic breast cancer. Targeted treatments for different subtypes of breast cancers have improved overall patient survival, even with metastasis. However, these targeted treatments are based upon the biology of the primary tumor and often these patients' relapse, after therapy, with metastatic tumors. The advent of immunotherapy allowed the immune system to target metastatic tumors. Unfortunately, immunotherapy has not been as effective in metastatic breast cancer relative to other cancers with metastases, such as melanoma. This review will describe the heterogeneic nature of breast cancer cells and their microenvironments. The distinct properties of metastatic breast cancer cells and their microenvironments that allow interactions, especially in bone and brain metastasis, will also be described. Finally, we will review immunotherapy approaches to treat metastatic breast tumors and discuss future therapeutic approaches to improve treatments for metastatic breast cancer.

The impact of COVID-19 on professional development for physiotherapists in Lagos, Nigeria.

Background: Continuous professional development is important to maintain standards of care in the healthcare sector. However, in Lagos, Nigeria, the additional burden of COVID-19 and limited resources may provide separate challenges for physiotherapists' continuous professional development (CPD). Objectives: To examine the availability and perceived quality of continuous professional development opportunities for physiotherapists working before and during the pandemic in Lagos, Nigeria. Method: A qualitative study was conducted with 10 conveniently sampled physiotherapists, recruited via email. Interviews took place via Zoom, and the video function was utilised. Data were collected via semi-structured interviews using a pilot tested interview, and was transcribed and analysed thematically. Results: The main method of workplace teaching pre-COVID-19 and during COVID-19 was bedside teaching (BT), which most participants received. Pre-COVID-19, the main barrier to receiving teaching was a lack of national guidelines providing specific details on CPD. During COVID-19, the main barrier was the difficulty of increased online teaching instead of teaching on real life patients. The main barriers for non-workplace CPD pre-COVID-19 were a lack of availability of learning opportunities and monetary cost of conferences. This was partially combatted by the increasing trend of online learning events during the pandemic, which increased access to non-workplace learning opportunities. Conclusion: Because of COVID-19, most CPD learning opportunities for physiotherapists in Lagos, Nigeria, were online, increasing overall accessibility. Adequate training to improve utilising online learning resources as well as specific guidelines for workplace physiotherapists CPD in Nigeria should be implemented and promoted to improve confidence and quality of care. Clinical implications: Key insight into the CPD experiences of physiotherapists currently working in Lagos, Nigeria, which can guide policies and improve clinical outcomes.

All-in-One digital microfluidics pipeline for proteomic sample preparation and analysis.

Highly sensitive and reproducible analysis of samples containing low amounts of protein is restricted by sample loss and the introduction of contaminants during processing. Here, we report an All-in-One digital microfluidic (DMF) pipeline for proteomic sample reduction, alkylation, digestion, isotopic labeling and analysis. The system features end-to-end automation, with integrated thermal control for digestion, optimized droplet additives for sample manipulation and analysis, and an automated interface to liquid chromatography with tandem mass spectrometry (HPLC-MS/MS). Dimethyl labeling was integrated into the pipeline to allow for relative quantification of the trace samples at the nanogram level, and the new pipeline was applied to evaluating cancer cell lines and cancer tissue samples. Several known proteins (including HSP90AB1, HSPB1, LDHA, ENO1, PGK1, KRT18, and AKR1C2) and pathways were observed between model breast cancer cell lines related to hormone response, cell metabolism, and cell morphology. Furthermore, differentially quantified proteins (such as PGS2, UGDH, ASPN, LUM, COEA1, and PRELP) were found in comparisons of healthy and cancer breast tissues, suggesting potential utility of the All-in-One pipeline for the emerging application of proteomic cancer sub-typing. In sum, the All-in-One pipeline represents a powerful new tool for automated proteome processing and analysis, with the potential to be useful for evaluating mass-limited samples for a wide range of applications.

Host parent responses to heterospecific parasite nestling alarm calls are independent of past and current experience with experimental brood parasitism.

Communication between parents and dependent offspring is critical not only during provisioning, but also in antipredator contexts. In altricial birds, a top cause of reproductive failure is nest predation, and alarm calls both by parents and chicks can serve to alert others and increase the likelihood of offspring escaping predation. Understanding the factors that determine the strength of parental antipredator responses to different nestling alarm calls can provide insight into parent-offspring recognition. The prothonotary warbler (Protonotaria citrea), a host of the obligate brood parasite, the brown-headed cowbird (Molothrus ater), never rejects cowbird young and raises the parasite together with its own offspring. To determine whether warbler parents learn cowbird nestling alarm calls, we presented experimentally parasitized or non-parasitized parents with playbacks of conspecific warbler, parasitic cowbird, and a harmless heterospecific control, eastern bluebird (Sialis sialis), nestling alarm calls. We recorded the latency to respond and the number of chips given by members of the resident warbler pair. We found that parents were most likely to respond to warbler nestling alarm calls, least likely to respond to bluebird calls, with a statistically intermediate likelihood of responding to cowbird calls. Critically, current and past parasitism status did not affect the likelihood of response to any playback or the number of chips given, however, currently parasitized parents had greater response latencies to playbacks than non-parasitized parents. These results suggest that warbler parents do not learn cowbird alarm calls from breeding experiences and, in turn, that cowbirds may employ a generalized, bet-hedging alarm call.

Physiotherapy in Madagascar: current challenges and opportunities for development.

PURPOSE: World Health Organisation's data show that low and middle-income countries have a higher prevalence of disabilities. Madagascar is the ninth poorest country in the world. This report aims to analyse the current difficulties faced by physiotherapists and physicians working in the hospital setting, and offer recommendations for how healthcare services can develop in the future. MATERIALS AND METHODS: Data were collected over the course of 3 months in the form of observational logs, interviews, and questionnaires. Interpreters were used in all interviews, and written questionnaires were translated from English to Malagasy. RESULTS: Thematic analysis of all data collected was completed with key themes emerging: difficult working conditions, including high patient volume and limited staff capacity or treatment space; limited recognition of physiotherapy as a profession, with no autonomy for physiotherapy staff; the low standard of clinical reasoning skills being used in practice; and variable levels of clinical competence, with little evidence of active continuing professional development or appropriate training specific to the needs of clinicians. CONCLUSIONS: The current practice of physiotherapy in Madagascar mirrors the challenge in low-income settings. This study highlights the ongoing needs of the profession, including increasing clinical reasoning skills, updating the physiotherapy curriculum, and further development of multidisciplinary teamwork.IMPLICATIONS FOR REHABILITATIONThere are approximately 300 physiotherapists in Madagascar serving 25.5 million people in a low-income country with a high prevalence of disability.Developing the core training skills of physiotherapists in areas such as clinical reasoning skills, improved multidisciplinary team working, and access to research will allow physiotherapists to deliver a higher level of patient-centred care focused on rehabilitation.Upskilling physiotherapists and improving rehabilitation standards will result in improved functioning and quality of life for people with disabilities.Developments in multidisciplinary team communication and working will lead to interprofessional respect, improved quality of work, and a focus on patient-centred care.

Intrafractional stability of MR-guided online adaptive SBRT for prostate cancer.

BACKGROUND: MR-guided online adaptive stereotactic body radiation therapy (SBRT) for prostate cancer aims to reduce toxicity by full compensation of interfractional uncertainties. However, the process of online adaptation currently takes approximately 45 min during which intrafractional movements remain unaccounted for. This study aims to analyze the dosimetric benefit of online adaptation and to evaluate its robustness over the duration of one treatment fraction. METHODS: Baseline MR-scans at a MR-linear accelerator were acquired for ten healthy male volunteers for generation of mock-prostate SBRT plans with a dose prescription of 5 × 7.25 Gy. On a separate day, online MR-guided adaptation (ViewRay® MRIdian) was performed, and thereafter MR images were acquired every 15 min for 1 h to assess the stability of the adapted plan. RESULTS: A dosimetric benefit of online MR-guided adaptive re-planning was observed in 90% of volunteers. The median D95CTV- and D95PTV-coverage was improved from 34.8 to 35.5 Gy and from 30.7 to 34.6 Gy, respectively. Improved target coverage was not associated with higher dose to the organs at risk, most importantly the rectum (median D1ccrectum baseline plan vs. adapted plan 33.3 Gy vs. 32.3 Gy). The benefit of online adaptation remained stable over 45 min for all volunteers. However, at 60 min, CTV-coverage was below a threshold of 32.5 Gy in 30% of volunteers (30.6 Gy, 32.0 Gy, 32.3 Gy). CONCLUSION: The dosimetric benefit of MR-guided online adaptation for prostate SBRT was robust over 45 min in all volunteers. However, intrafractional uncertainties became dosimetrically relevant at 60 min and we therefore recommend verification imaging before delivery of MR-guided online adapted SBRT.

Reconfigurable multi-component micromachines driven by optoelectronic tweezers.

There is great interest in the development of micromotors which can convert energy to motion in sub-millimeter dimensions. Micromachines take the micromotor concept a step further, comprising complex systems in which multiple components work in concert to effectively realize complex mechanical tasks. Here we introduce light-driven micromotors and micromachines that rely on optoelectronic tweezers (OET). Using a circular micro-gear as a unit component, we demonstrate a range of new functionalities, including a touchless micro-feed-roller that allows the programming of precise three-dimensional particle trajectories, multi-component micro-gear trains that serve as torque- or velocity-amplifiers, and micro-rack-and-pinion systems that serve as microfluidic valves. These sophisticated systems suggest great potential for complex micromachines in the future, for application in microrobotics, micromanipulation, microfluidics, and beyond.

Delays to accessing healthcare and rehabilitation following trauma in Madagascar - a qualitative study.

PURPOSE: To explore the factors affecting access to timely trauma care and rehabilitation in Madagascar. MATERIALS AND METHODS: A qualitative study based in the outpatient departments of two large rehabilitation centres. Semi-structured interviews and focus groups were conducted with 12 patients or family members and 11 healthcare professionals. Interviews and focus groups were conducted with a local interpreter and were audio-recorded and transcribed. The data were analysed deductively with thematic content analysis, utilising the Health Care Access Barriers model. RESULTS: Participants experienced delays in deciding to seek treatment, accessing healthcare facilities and in receiving appropriate treatment. Cognitive barriers included understanding and awareness of healthcare, structural barriers included distance and transportation to health facilities, financial barriers included affordability of healthcare and difficulty accessing funds. CONCLUSIONS: Delays to accessing healthcare may result in increased mortality and disability following trauma, as well as increased financial burden. Addressing the acceptability of services should be a focus for future service development, through training and education schemes. More importantly, improving both physical and financial accessibility of services must be a long-term priority. These findings may help to guide the ongoing development of trauma and rehabilitation pathways in Madagascar.IMPLICATIONS FOR REHABILITATIONDelays in accessing timely trauma care and rehabilitation can lead to complications such as pain, infection, malunion of fractures and loss of function, with devastating financial and social consequences for patients and their families.The acceptability of services is a key barrier to accessing hospital care and may be targeted through training schemes for healthcare professionals and education for the public.Improving the physical and financial accessibility of services in the long-term is paramount.Consideration should be given to these issues in the future development of co-ordinated trauma care and rehabilitation pathways in Madagascar.

Digital microfluidic isolation of single cells for -Omics.

We introduce Digital microfluidic Isolation of Single Cells for -Omics (DISCO), a platform that allows users to select particular cells of interest from a limited initial sample size and connects single-cell sequencing data to their immunofluorescence-based phenotypes. Specifically, DISCO combines digital microfluidics, laser cell lysis, and artificial intelligence-driven image processing to collect the contents of single cells from heterogeneous populations, followed by analysis of single-cell genomes and transcriptomes by next-generation sequencing, and proteomes by nanoflow liquid chromatography and tandem mass spectrometry. The results described herein confirm the utility of DISCO for sequencing at levels that are equivalent to or enhanced relative to the state of the art, capable of identifying features at the level of single nucleotide variations. The unique levels of selectivity, context, and accountability of DISCO suggest potential utility for deep analysis of any rare cell population with contextual dependencies.

Cell invasion in digital microfluidic microgel systems.

Microfluidic methods for studying cell invasion can be subdivided into those in which cells invade into free space and those in which cells invade into hydrogels. The former techniques allow straightforward extraction of subpopulations of cells for RNA sequencing, while the latter preserve key aspects of cell interactions with the extracellular matrix (ECM). Here, we introduce "cell invasion in digital microfluidic microgel systems" (CIMMS), which bridges the gap between them, allowing the stratification of cells on the basis of their invasiveness into hydrogels for RNA sequencing. In initial studies with a breast cancer model, 244 genes were found to be differentially expressed between invading and noninvading cells, including genes correlating with ECM-remodeling, chemokine/cytokine receptors, and G protein transducers. These results suggest that CIMMS will be a valuable tool for probing metastasis as well as the many physiological processes that rely on invasion, such as tissue development, repair, and protection.

Trauma rehabilitation in a teaching hospital in Antananarivo, Madagascar: current provision and patients' perspectives.

Purpose: This study aims to describe the current provision of trauma rehabilitation in Antananarivo, Madagascar and explore the experiences and perspectives of users of this service in order to inform future research and service development.Material and methods: A qualitative study supplemented by a descriptive service review were applied in the central government hospital in Antananarivo. Registers in the rehabilitation department were analyzed for a 3 month period. Semi-structured interviews were conducted with a convenience sample of users of the rehabilitation service and were subject to thematic analysis.Results: One in six patients treated in the rehabilitation department has survived a traumatic injury, with limb injuries comprising the majority. Analysis of interviews with 10 adult patients, following traumatic injuries to the lower limbs (6), upper limbs (3) or polytrauma (1) generated seven broad themes: health seeking behavior, organization of services, clinical management, costs and financial impact, effect on function and role, psychological impact and societal attitudes.Conclusions: Financial constraints and impact on personal finances pervade the overall picture. The perspectives and experiences elucidated will inform future research into the development of context-specific models of care for people with trauma-related disabilities in Madagascar.Implications for rehabilitationPotentially treatable traumatic injury can have a profound effect on function and societal participation in Madagascar.Interventions to increase access to timely acute trauma care and rehabilitation are needed, and may include different models of financing and innovations to reduce materials costs and length of stay.A structured rehabilitation pathway must be integrated to the earliest stages of trauma care, including scrutinizing the use of prolonged immobilization.Consideration should be given to models of care which are financially and geographically accessible to the population.

Evaluating a global health partnership rehabilitation training programme in Madagascar.

OBJECTIVE: Rehabilitation services play an important role in optimizing functional ability and societal integration for people with disabilities. The Madagascar Rehabilitation Programme (2011-2013) resulted from a global training partnership and led to 8 doctors achieving a university diploma in rehabilitation medicine. This paper describes a 2014 evaluation of the programme methods, results and learning points. METHODS: A combination of qualitative methods was used for the evaluation, based on a Theory of Change model, with informants from Madagascar and the UK. RESULTS: Malagasy trainees and UK volunteers gained new theoretical knowledge and practical skills. For Madagascar, it led to changes in working practice and the formation of a national rehabilitation association. Key to its success was the strong collaboration between Malagasy and UK professionals, with support from the University and Ministry of Health in Madagascar, and the UK partners. Having a clear common vision ensured the programme met the needs of the Malagasy clinicians. CONCLUSION: Rehabilitation is increasingly recognized as an important focus for international development. Successful rehabilitation training programmes can be achieved at modest costs with global health partnerships. The combination of factors that enabled this programme to be a success is reproducible in other contexts.

The optoelectronic microrobot: A versatile toolbox for micromanipulation.

Microrobotics extends the reach of human-controlled machines to submillimeter dimensions. We introduce a microrobot that relies on optoelectronic tweezers (OET) that is straightforward to manufacture, can take nearly any desirable shape or form, and can be programmed to carry out sophisticated, multiaxis operations. One particularly useful program is a serial combination of "load," "transport," and "deliver," which can be applied to manipulate a wide range of micrometer-dimension payloads. Importantly, microrobots programmed in this manner are much gentler on fragile mammalian cells than conventional OET techniques. The microrobotic system described here was demonstrated to be useful for single-cell isolation, clonal expansion, RNA sequencing, manipulation within enclosed systems, controlling cell-cell interactions, and isolating precious microtissues from heterogeneous mixtures. We propose that the optoelectronic microrobotic system, which can be implemented using a microscope and consumer-grade optical projector, will be useful for a wide range of applications in the life sciences and beyond.

Patterned Optoelectronic Tweezers: A New Scheme for Selecting, Moving, and Storing Dielectric Particles and Cells.

Optical micromanipulation has become popular for a wide range of applications. In this work, a new type of optical micromanipulation platform, patterned optoelectronic tweezers (p-OET), is introduced. In p-OET devices, the photoconductive layer (that is continuous in a conventional OET device) is patterned, forming regions in which the electrode layer is locally exposed. It is demonstrated that micropatterns in the photoconductive layer are useful for repelling unwanted particles/cells, and also for keeping selected particles/cells in place after turning off the light source, minimizing light-induced heating. To clarify the physical mechanism behind these effects, systematic simulations are carried out, which indicate the existence of strong nonuniform electric fields at the boundary of micropatterns. The simulations are consistent with experimental observations, which are explored for a wide variety of geometries and conditions. It is proposed that the new technique may be useful for myriad applications in the rapidly growing area of optical micromanipulation.

Southern Hemisphere westerlies as a driver of the early deglacial atmospheric CO2 rise.

The early part of the last deglaciation is characterised by a ~40 ppm atmospheric CO2 rise occurring in two abrupt phases. The underlying mechanisms driving these increases remain a subject of intense debate. Here, we successfully reproduce changes in CO2, δ13C and Δ14C as recorded by paleo-records during Heinrich stadial 1 (HS1). We show that HS1 CO2 increase can be explained by enhanced Southern Ocean upwelling of carbon-rich Pacific deep and intermediate waters, resulting from intensified Southern Ocean convection and Southern Hemisphere (SH) westerlies. While enhanced Antarctic Bottom Water formation leads to a millennial CO2 outgassing, intensified SH westerlies induce a multi-decadal atmospheric CO2 rise. A strengthening of SH westerlies in a global eddy-permitting ocean model further supports a multi-decadal CO2 outgassing from the Southern Ocean. Our results highlight the crucial role of SH westerlies in the global climate and carbon cycle system with important implications for future climate projections.

A digital microfluidic system for serological immunoassays in remote settings.

Serosurveys are useful for assessing population susceptibility to vaccine-preventable disease outbreaks. Although at-risk populations in remote areas could benefit from this type of information, they face several logistical barriers to implementation, such as lack of access to centralized laboratories, cold storage, and transport of samples. We describe a potential solution: a compact and portable, field-deployable, point-of-care system relying on digital microfluidics that can rapidly test a small volume of capillary blood for disease-specific antibodies. This system uses inexpensive, inkjet-printed digital microfluidic cartridges together with an integrated instrument to perform enzyme-linked immunosorbent assays (ELISAs). We performed a field validation of the system's analytical performance at Kakuma refugee camp, a remote setting in northwestern Kenya, where we tested children aged 9 to 59 months and caregivers for measles and rubella immunoglobulin G (IgG). The IgG assays were determined to have sensitivities of 86% [95% confidence interval (CI), 79 to 91% (measles)] and 81% [95% CI, 73 to 88% (rubella)] and specificities of 80% [95% CI, 49 to 94% (measles)] and 91% [95% CI, 76 to 97% (rubella)] (measles, n = 140; rubella, n = 135) compared with reference tests (measles IgG and rubella IgG ELISAs from Siemens Enzygnost) conducted in a centralized laboratory. These results demonstrate a potential role for this point-of-care system in global serological surveillance, particularly in remote areas with limited access to centralized laboratories.

Escape from an Optoelectronic Tweezer Trap: experimental results and simulations.

Optoelectronic tweezers (OET) are a microsystem actuation technology capable of moving microparticles at mm s-1 velocities with nN forces. In this work, we analyze the behavior of particles manipulated by negative dielectrophoresis (DEP) forces in an OET trap. A user-friendly computer interface was developed to generate a circular rotating light pattern to control the movement of the particles, allowing their force profiles to be conveniently measured. Three-dimensional simulations were carried out to clarify the experimental results, and the DEP forces acting on the particles were simulated by integrating the Maxwell stress tensor. The simulations matched the experimental results and enabled the determination of a new "hopping" mechanism for particle-escape from the trap. As indicated by the simulations, there exists a vertical DEP force at the edge of the light pattern that pushes up particles to a region with a smaller horizontal DEP force. We propose that this phenomenon will be important to consider for the design of OET micromanipulation experiments for a wide range of applications.

Developing a spinal cord injury rehabilitation service in Madagascar.

Rehabilitation for people with spinal cord injury in many low- and middle-income countries is not avail-able or is in the early stages of development. However, rehabilitation is recognized as crucial in order to optimize functional recovery and outcomes for patients with spinal cord injury. With an increasing incidence of spinal cord injury, the unmet need for rehabilitation is huge. This report describes the early development of a specialist rehabilitation service for spinal cord injury in Madagascar, one of the poorest countries in the world. The sustained input to an expanding rehabilitation team has led to reductions in avoidable complications. The input of the rehabilitation team has been welcomed by the neurosurgery department, which has recognized fewer delays in patients undergoing surgical treatments. Cost, lack of resources and trained staff, and poor understanding of disability continue to provide challenges. However, the development of the rehabilitation service using low technology, but with a high level of knowledge and systematic management, is a source of considerable pride. This development in Madagascar can be regarded as a model for spinal cord injury rehabilitation in other low-resource settings.

Pre-concentration by liquid intake by paper (P-CLIP): a new technique for large volumes and digital microfluidics.

Microfluidic platforms are an attractive option for incorporating complex fluid handling into low-cost and rapid diagnostic tests. A persistent challenge for microfluidics, however, is the mismatch in the "world-to-chip" interface - it is challenging to detect analytes present at low concentrations in systems that can only handle small volumes of sample. Here we describe a new technique termed pre-concentration by liquid intake by paper (P-CLIP) that addresses this mismatch, allowing digital microfluidics to interface with volumes on the order of hundreds of microliters. In P-CLIP, a virtual microchannel is generated to pass a large volume through the device; analytes captured on magnetic particles can be isolated and then resuspended into smaller volumes for further processing and analysis. We characterize this method and demonstrate its utility with an immunoassay for Plasmodium falciparum lactate dehydrogenase, a malaria biomarker, and propose that the P-CLIP strategy may be useful for a wide range of applications that are currently limited by low-abundance analytes.

The use of cell culture platforms to identify novel markers of bone and dentin resorption.

OBJECTIVES: 1) To test the hypothesis that there would be proteomic differences in the composition of exosomes isolated from osteoclasts and odontoclasts and 2) to determine the clinical usefulness of these in vitro biomarker candidates. MATERIALS AND METHODS: Mouse bone marrow-derived precursors were cultured on either dentin or bone slices and allowed to mature and begin resorption. Exosomes were isolated from cell culture media and characterized by mass spectrometry. The proteomic data obtained from this in vitro study were compared with the data obtained from human samples in our previous work. RESULTS: There was a difference in the proteomic composition of exosomes from osteoclasts and odontoclasts. A total of 40 exosomal proteins were only present in osteoclast media, whereas six unique exosomal proteins were identified in odontoclast supernatants. Approximately 50% of exosomal proteins released by clastic cells in vitro can be found in oral fluids. CONCLUSION: Our data suggest that the mineralized matrix type plays a role in the final phenotypic characteristics of mouse clastic cells. Many in vitro biomarker candidates of bone and dentin resorption can also be found in human oral fluids, thus indicating that this approach may be a viable alternative in biomarker discovery.