Platelet lysates from aged donors promote human tenocyte proliferation and migration in a concentration-dependent manner.

OBJECTIVES: Platelet-rich plasma (PRP) is being used increasingly often in the clinical setting to treat tendon-related pathologies. Yet the optimal PRP preparations to promote tendon healing in different patient populations are poorly defined. Here, we sought to determine whether increasing the concentration of platelet-derived proteins within a derivative of PRP, platelet lysate (PL), enhances tenocyte proliferation and migration in vitro, and whether the mitogenic properties of PL change with donor age. METHODS: Concentrated PLs from both young (< 50 years) and aged (> 50 years) donors were prepared by exposing pooled PRP to a series of freeze-thaw cycles followed by dilution in plasma, and the levels of several platelet-derived proteins were measured using multiplex immunoassay technology. Human tenocytes were cultured with PLs to simulate a clinically relevant PRP treatment range, and cell growth and migration were assessed using DNA quantitation and gap closure assays, respectively. RESULTS: Platelet-derived protein levels increased alongside higher PL concentrations, and PLs from both age groups improved tenocyte proliferation relative to control conditions. However, PLs from aged donors yielded a dose-response relationship in tenocyte behaviour, with higher PL concentrations resulting in increased tenocyte proliferation and migration. Conversely, no significant differences in tenocyte behaviour were detected when increasing the concentration of PLs from younger donors. CONCLUSION: Higher PL concentrations, when prepared from the PRP of aged but not young donors, were more effective than lower PL concentrations at promoting tenocyte proliferation and migration in vitro.Cite this article: D. R. Berger, C. J. Centeno, N. J. Steinmetz. Platelet lysates from aged donors promote human tenocyte proliferation and migration in a concentration-dependent manner. Bone Joint Res 2019;8:32-40. DOI: 10.1302/2046-3758.81.BJR-2018-0164.R1.

Cancer Theranostic Applications of Albumin-Coated Tobacco Mosaic Virus Nanoparticles.

Nanotechnology holds great promise in cancer drug delivery, and of particular interest are theranostic approaches in which drug delivery and imaging are integrated. In this work, we studied and developed the plant virus tobacco mosaic virus (TMV) as a platform nanotechnology for drug delivery and imaging. Specifically, a serum albumin (SA)-coated TMV formulation was produced. The SA coating fulfils two functions: SA provides a stealth coating for enhanced biocompatibility; it also acts as a targeting ligand enabling efficient tumor accumulation of SA-TMV versus TMV in mouse models of breast and prostate cancer. We demonstrate drug delivery of the chemotherapy doxorubicin (DOX); TMV-delivered DOX outperformed free DOX, resulting in significant delayed tumor growth and increased survival. Furthermore, we demonstrated the ability of SA-coated TMV loaded with chelated Gd(DOTA) for magnetic resonance imaging detection of tumors. In the future, we envision the application of such probes as theranostic, where first imaging is performed to assess whether the nanoparticles are effective at targeting a particular patient tumor. If targeting is confirmed, the therapeutic would be added and treatment can begin. The combination of imaging and therapy would allow to monitor disease progression and therefore inform about the effectiveness of the drug delivery approach.

The in vivo fates of plant viral nanoparticles camouflaged using self-proteins: overcoming immune recognition.

Nanoparticles offer a promising avenue for targeted delivery of therapies. To slow clearance, nanoparticles are frequently stealth-coated to prevent opsonization and immune recognition. Serum albumin (SA) has been used as a bio-inspired stealth coating. To develop this shielding strategy for clinical applications, it is critical to understand the interactions between the immune system and SA-camouflaged nanoparticles. This work investigates the in vivo processing of SA-coated nanoparticles using tobacco mosaic virus (TMV) as a model system. In comparing four different SA-formulations, the particles with high SA coverage conjugated to TMV via a short linker performed the best at preventing antibody recognition. Irrelevant of the coating chemistry, all formulations led to similar levels of TMV-specific antibodies after repeat administration in mice; importantly though, SA-specific antibodies were not detected and the TMV-specific antibodies were unable to recognize shielded SA-coated TMV. Upon uptake in macrophages, the shielding agent and nanoparticle separate, where TMV trafficked to the lysosome and SA appears to recycle. The distinct intracellular fates of the TMV carrier and SA shielding agent explain why anti-TMV but not SA-specific antibodies are generated. This work characterizes the outcomes of SA-camouflaged TMV after immune recognition, and highlights the effectiveness of SA as a nanoparticle shielding agent.

In situ vaccination with cowpea mosaic virus nanoparticles suppresses metastatic cancer.

Nanotechnology has tremendous potential to contribute to cancer immunotherapy. The 'in situ vaccination' immunotherapy strategy directly manipulates identified tumours to overcome local tumour-mediated immunosuppression and subsequently stimulates systemic antitumour immunity to treat metastases. We show that inhalation of self-assembling virus-like nanoparticles from cowpea mosaic virus (CPMV) reduces established B16F10 lung melanoma and simultaneously generates potent systemic antitumour immunity against poorly immunogenic B16F10 in the skin. Full efficacy required Il-12, Ifn-γ, adaptive immunity and neutrophils. Inhaled CPMV nanoparticles were rapidly taken up by and activated neutrophils in the tumour microenvironment as an important part of the antitumour immune response. CPMV also exhibited clear treatment efficacy and systemic antitumour immunity in ovarian, colon, and breast tumour models in multiple anatomic locations. CPMV nanoparticles are stable, nontoxic, modifiable with drugs and antigens, and their nanomanufacture is highly scalable. These properties, combined with their inherent immunogenicity and demonstrated efficacy against a poorly immunogenic tumour, make CPMV an attractive and novel immunotherapy against metastatic cancer.

Molecular farming of fluorescent virus-based nanoparticles for optical imaging in plants, human cells and mouse models.

The application of plant virus-derived nanostructures in materials science, biomedical research and engineering has recently been promoted by the development of fluorescence-labeled viruses for optical imaging in tissue culture and preclinical animal models. Most studies reported thus far have focused on the application of viruses that have been chemically modified with organic dyes. In this investigation, we sought to develop and study genetically-engineered virus-based biomaterials that incorporate green or red fluorescent proteins. The genetic introduction of such imaging moieties is advantageous because post-harvest modifications are not required, thus minimizing the number of manufacturing steps and maximizing the yields of each fluorescent probe. Specifically, we engineered the filamentous plant virus Potato virus X (PVX) to display green fluorescent protein (GFP) or mCherry as N-terminal coat protein (CP) fusions, producing a 1 : 3 fusion protein to CP ratio. The infection of Nicotiana benthamiana plants with the recombinant GFP-PVX and mCherry-PVX particles was documented by fluorescence imaging, structural analysis and genetic characterization to determine the stability of the chimeras and optimize the molecular farming protocols. We also demonstrated the application of fluorescent mCherry-PVX filaments as probes for optical imaging in human cancer cells and a preclinical mouse model. Cell viability assays and histological analysis following the administration of mCherry-PVX indicated the biocompatibility and rapid tissue clearance of the particles. Such particles could therefore be functionalized with additional cancer-specific detection ligands to provide tools for molecular imaging, allowing the investigation of molecular signatures, disease progression/recurrence and the efficacy of novel therapies.

Structure-based engineering of an icosahedral virus for nanomedicine and nanotechnology.

A quintessential tenet of nanotechnology is the self-assembly of nanometer-sized components into devices. Biological macromolecular systems such as viral particles were found to be suitable building blocks for nanotechnology for several reasons: viral capsids are extremely robust and can be produced in large quantities with ease, the particles self-assemble into monodisperse particles with a high degree of symmetry and polyvalency, they have the propensity to form arrays, and they offer programmability through genetic and chemical engineering. Here, we review the recent advances in engineering the icosahedral plant virus Cowpea mosaic virus (CPMV) for applications in nano-medicine and -technology. In the first part, we will discuss how the combined knowledge of the structure of CPMV at atomic resolution and the use of chimeric virus technology led to the generation of CPMV particles with short antigenic peptides for potential use as vaccine candidates. The second part focuses on the chemical addressability of CPMV. Strategies to chemically attach functional molecules at designed positions on the exterior surface of the viral particle are described. Biochemical conjugation methods led to the fabrication of electronically conducting CPMV particles and networks. In addition, functional proteins for targeted delivery to mammalian cells were successfully attached to CPMV. In the third part, we focus on the utilization of CPMV as a building block for the generation of 2D and 3D arrays. Overall, the potential applications of viral nanobuilding blocks are manifold and range from nanoelectronics to biomedical applications.

Comparison of forced-air warming systems with lower body blankets using a copper manikin of the human body.

BACKGROUND: Forced-air warming has gained high acceptance as a measure for the prevention of intraoperative hypothermia. However, data on heat transfer with lower body blankets are not yet available. This study was conducted to determine the heat transfer efficacy of six complete lower body warming systems. METHODS: Heat transfer of forced-air warmers can be described as follows:[1]Qdot;=h.DeltaT.A where Qdot; = heat transfer [W], h = heat exchange coefficient [W m-2 degrees C-1], DeltaT = temperature gradient between blanket and surface [ degrees C], A = covered area [m2]. We tested the following forced-air warmers in a previously validated copper manikin of the human body: (1) Bair Hugger and lower body blanket (Augustine Medical Inc., Eden Prairie, MN); (2) Thermacare and lower body blanket (Gaymar Industries, Orchard Park, NY); (3) WarmAir and lower body blanket (Cincinnati Sub-Zero Products, Cincinnati, OH); (4) Warm-Gard(R) and lower body blanket (Luis Gibeck AB, Upplands Väsby, Sweden); (5) Warm-Gard and reusable lower body blanket (Luis Gibeck AB); and (6) WarmTouch and lower body blanket (Mallinckrodt Medical Inc., St. Luis, MO). Heat flux and surface temperature were measured with 16 calibrated heat flux transducers. Blanket temperature was measured using 16 thermocouples. DeltaT was varied between -10 and +10 degrees C and h was determined by a linear regression analysis as the slope of DeltaT vs. heat flux. Mean DeltaT was determined for surface temperatures between 36 and 38 degrees C, because similar mean skin temperatures have been found in volunteers. The area covered by the blankets was estimated to be 0.54 m2. RESULTS: Heat transfer from the blanket to the manikin was different for surface temperatures between 36 degrees C and 38 degrees C. At a surface temperature of 36 degrees C the heat transfer was higher (between 13.4 W to 18.3 W) than at surface temperatures of 38 degrees C (8-11.5 W). The highest heat transfer was delivered by the Thermacare system (8.3-18.3 W), the lowest heat transfer was delivered by the Warm-Gard system with the single use blanket (8-13.4 W). The heat exchange coefficient varied between 12.5 W m-2 degrees C-1 and 30.8 W m-2 degrees C-1, mean DeltaT varied between 1.04 degrees C and 2.48 degrees C for surface temperatures of 36 degrees C and between 0.50 degrees C and 1.63 degrees C for surface temperatures of 38 degrees C. CONCLUSION: No relevant differences in heat transfer of lower body blankets were found between the different forced-air warming systems tested. Heat transfer was lower than heat transfer by upper body blankets tested in a previous study. However, forced-air warming systems with lower body blankets are still more effective than forced-air warming systems with upper body blankets in the prevention of perioperative hypothermia, because they cover a larger area of the body surface.

Comparison of forced-air warming systems with upper body blankets using a copper manikin of the human body.

BACKGROUND: Forced-air warming with upper body blankets has gained high acceptance as a measure for the prevention of intraoperative hypothermia. However, data on heat transfer with upper body blankets are not yet available. This study was conducted to determine the heat transfer efficacy of eight complete upper body warming systems and to gain more insight into the principles of forced-air warming. METHODS: Heat transfer of forced-air warmers can be described as follows: Qdot;=h. DeltaT. A, where Qdot;= heat flux [W], h=heat exchange coefficient [W m-2 degrees C-1], DeltaT=temperature gradient between the blanket and surface [ degrees C], and A=covered area [m2]. We tested eight different forced-air warming systems: (1) Bair Hugger and upper body blanket (Augustine Medical Inc. Eden Prairie, MN); (2) Thermacare and upper body blanket (Gaymar Industries, Orchard Park, NY); (3) Thermacare (Gaymar Industries) with reusable Optisan upper body blanket (Willy Rüsch AG, Kernen, Germany); (4) WarmAir and upper body blanket (Cincinnati Sub-Zero Products, Cincinnati, OH); (5) Warm-Gard and single use upper body blanket (Luis Gibeck AB, Upplands Väsby, Sweden); (6) Warm-Gard and reusable upper body blanket (Luis Gibeck AB); (7) WarmTouch and CareDrape upper body blanket (Mallinckrodt Medical Inc., St. Luis, MO); and (8) WarmTouch and reusable MultiCover trade mark upper body blanket (Mallinckrodt Medical Inc.) on a previously validated copper manikin of the human body. Heat flux and surface temperature were measured with 11 calibrated heat flux transducers. Blanket temperature was measured using 11 thermocouples. The temperature gradient between the blanket and surface (DeltaT) was varied between -8 and +8 degrees C, and h was determined by linear regression analysis as the slope of DeltaT vs. heat flux. Mean DeltaT was determined for surface temperatures between 36 and 38 degrees C, as similar mean skin surface temperatures have been found in volunteers. The covered area was estimated to be 0.35 m2. RESULTS: Total heat flow from the blanket to the manikin was different for surface temperatures between 36 and 38 degrees C. At a surface temperature of 36 degrees C the heat flows were higher (4-26.6 W) than at surface temperatures of 38 degrees C (2.6-18.1 W). The highest total heat flow was delivered by the WarmTouch trade mark system with the CareDrape trade mark upper body blanket (18.1-26.6 W). The lowest total heat flow was delivered by the Warm-Gard system with the single use upper body blanket (2.6-4 W). The heat exchange coefficient varied between 15.1 and 36.2 W m-2 degrees C-1, and mean DeltaT varied between 0.5 and 3.3 degrees C. CONCLUSION: We found total heat flows of 2.6-26.6 W by forced-air warming systems with upper body blankets. However, the changes in heat balance by forced-air warming systems with upper body blankets are larger, as these systems are not only transferring heat to the body but are also reducing heat losses from the covered area to zero. Converting heat losses of approximately 37.8 W to heat gain, results in a 40.4-64.4 W change in heat balance. The differences between the systems result from different heat exchange coefficients and different mean temperature gradients. However, the combination of a high heat exchange coefficient with a high mean temperature gradient is rare. This fact offers some possibility to improve these systems.

Annexin-V imaging for noninvasive detection of cardiac allograft rejection.

Heart transplant rejection is characterized pathologically by myocyte necrosis and apoptosis associated with interstitial mononuclear cell infiltration. Any one of these components can be targeted for noninvasive detection of transplant rejection. During apoptotic cell death, phosphatidylserine, a phospholipid that is normally confined to the inner leaflet of cell membrane bilayer, gets exteriorized. Technetium-99m-labeled annexin-V, an endogenous protein that has high affinity for binding to phosphatidylserine, has been administered intravenously for noninvasive identification of apoptotic cell death. In the present study of 18 cardiac allograft recipients, 13 patients had negative and five had positive myocardial uptake of annexin. These latter five demonstrated at least moderate transplant rejection and caspase-3 staining, suggesting apoptosis in their biopsy specimens. This study reveals the clinical feasibility and safety of annexin-V imaging for noninvasive detection of transplant rejection by targeting cell membrane phospholipid alterations that are commonly associated with the process of apoptosis.

Health care in big cities--the McGill experience.

Big cities encounter problems which are peculiar to them: population mix, housing conditions, social structure, infrastructure including transport, etc, which manifest in hospitals. From the beginning of the 20th century, big cities have been served by many providers of hospital care. Common to most cities is the change in the principles and methods of care, e.g. home care, and the notion that traditional hospital services are no longer the only answer to the needs of the population of large cities. Over the last decade, plans and projects have been devised to address the problems of excessive bed capacity, insufficient and excessive specialised units and difficulties in administration of payment systems.

Forecasting patient services: a 21st century vision of an academic health centre.

This article provides an overview of the development and implementation of the McGill University Health Centre model for forecasting patient services in the year 2004, and advice on how to apply the model. Critical success factors and case examples are highlighted. The insights provided will be of value to hospitals and other institutions that recognize the necessity of engaging in long-range planning and forecasting.

Legal issues in human clinical investigation: a primer for physicians.

Human clinical investigation plays a central role in contemporary nuclear cardiology. It is a complex expensive endeavor triggering multiple responsibilities not otherwise encountered by physicians in clinical practice. Clinical investigators, drug and device manufacturers, hospitals and universities, governmental agencies, research subjects, patient advocacy groups, and investors all have a legitimate interest in the conduct and the outcome of clinical research. Past abuses of research subjects in many countries including those unknowingly exposed to radioactive isotopes have raised ethical concerns that are now addressed by overlapping laws, regulations, guidelines, and institutional policies that have evolved to guide the research enterprise. The policies underlying this legal and regulatory structure include protection of research subjects, protection of the integrity of research design and research data, and protection of the public and private purse. Knowledge of these laws and regulations is essential for physicians participating in clinical investigations to protect their professional and personal interests. This article reviews legal issues relevant to physicians conducting human clinical trials in the United States.

Detection of wooden foreign bodies in muscle tissue: experimental comparison of computed tomography, magnetic resonance imaging, and ultrasonography.

An experimental study was performed to compare computed tomography (CT), magnetic resonance imaging (MRI), and real-time, high resolution ultrasonography (US) for the detection of wooden foreign bodies in muscle tissue. Wooden splinters were prepared, measured for size, soaked in saline, and placed in porcine muscle distant from and adjacent to bone. The specimens were then examined using roentgenography, CT and MRI in planes parallel and perpendicular to the splinters, and US. The largest wooden foreign bodies (minimum smallest width = 10 mm) were easily detected by CT, MRI, and US. Almost all splinters of various sizes, small and large, soaked in saline for 5 months were easily detected by MRI. Smaller splinters (minimum smallest width = 1-4 mm) soaked for only 3 days and placed distant from bone were most easily detected by US; those soaked for 5 months were most easily detected by either US or MRI. The smaller splinters soaked for only 3 days and placed near bone were not reliably detected by any of the imaging methods; CT and MRI were both more sensitive than US in this situation. MRI scanning was more sensitive perpendicular than parallel to the long axis of the splinters. Therefore, either US or MRI may be the best initial imaging modality for evaluation of a suspected wooden foreign body, depending on availability of imaging method, chronicity of symptoms, and proximity to bone.

A study of mortality near sour gas refineries in southwest Alberta: an epidemic unrevealed.

Concerns about excesses in a wide array of adverse health outcomes have been expressed for over 25 years by a rural population in southwestern Alberta, living downwind from natural gas refineries. Among these has been the perception that deaths have occurred more frequently than ought to have been expected. As part of a large field epidemiologic study undertaken during the summer of 1985 to investigate possible health effects in this area, a residential cohort study was carried out to study mortality. The cohort was defined as all those individuals who resided in the area in 1970. A total of 30,175 person-years of risk within Alberta were experienced by this cohort during 1970-84. The deaths during this period were enumerated by resident reports and by manual record linkage with the death records of the Alberta Bureau of Vital Statistics. Age- and sex-standardized mortality ratios, based on expected rates from 2 pre-specified demographically similar, non-metropolitan Southern Alberta populations, were 0.88 and 0.84 respectively, neither of which was significantly different from unity. These data cannot address the question of etiology but they can do much to allay the anxieties of a community convinced it had experienced an epidemic of death.

Chronic exposure to sour gas emissions: meeting a community concern with epidemiologic evidence.

For 25 years residents of a rural area in southwestern Alberta have complained of health problems attributed to sour gas emissions from nearby natural gas refineries. We undertook a large epidemiologic study of the current health status and the selected morbidity rates among 2152 people in the exposed area. We established two comparison groups: one was a demographically similar unexposed population and the other a demographically different group also exposed to sour gas emissions in another region. The methods included a cross-sectional survey of current residents and separate historical cohort studies involving registry linkage to investigate cancer incidence and all-cause mortality. The cross-sectional survey involved a comprehensive health questionnaire, standardized clinical examinations by physicians blinded to the subjects' symptoms and concerns, and several laboratory tests. We were able to contact just under 60% of the people who we knew had moved from each area since 1958 and found no evidence of selective migration for health reasons. Although the residents of the exposed area reported an excess number of symptoms and health problems there were no significant differences in the mortality rate, incidence of cancer, reproductive problems, major ailments, hair levels of arsenic and certain metals or respiratory function between the groups.

Respiratory health of a population living downwind from natural gas refineries.

Since 1958 there has been a perception of excess illness in a rural Canadian population living downwind from two natural gas refineries, the emissions of which contain mostly sulfur dioxide but also hydrogen sulfide. To determine if there was an excess of adverse health outcomes in the population exposed (defined by place of residence), a health survey was undertaken in 1985 in this area and in one unexposed to emissions but demographically similar. Participation was 92% from both the exposed population (n = 2,157) and a representative sample (n = 839) of the main reference population. More respiratory symptoms were reported in the exposed group than in the non-exposed group among those 5 to 13 yrs of age (28% versus 18%) and among never-smokers greater than or equal to 14 yrs of age (35% versus 24%). FEV1, FVC, and FEV1/FVC were similar in both areas. Dichotomizing the level of exposure (high, low) within the exposed area revealed a trend in the high exposure area toward increased respiratory symptoms in the younger age group (39% versus 24%), but decreased symptoms in the older age group (33% versus 36% among never-smokers). FEV1 was similar between the two areas. The excess of respiratory symptoms in the exposed area unassociated with impaired spirometric values would be compatible with increased awareness of health or a small biologic environmental effect.

Cancer downwind from sour gas refineries: the perception and the reality of an epidemic.

A rural population in southwestern Alberta, Canada, living downwind from natural gas refineries, has expressed concerns about an excess of adverse health outcomes over the last 25 years. This has escalated to the point of causing a prominent sociopolitical controversy within the province. As part of a large field epidemiologic study undertaken during the summer of 1985 to investigate possible health effects, a residential cohort study was carried out to study cancer incidence. The cohort was defined as all those individuals who resided in the area in 1970. A total of 30,175 person-years of risk within Alberta were experienced by this cohort from 1970 to 1984. The incident cancers during this period were enumerated by computerized record linkage with the Alberta Cancer Registry. Age- and sex-standardized incidence ratios, based on expected rates from three prespecified demographically similar, nonmetropolitan Southern Alberta populations, were 1.05, 1.09, and 1.03, respectively, none of which was significantly different from unity. Although they do not address the issue of etiologic association, these data can provide considerable reassurance to a community that was convinced it had experienced an epidemic of cancer.

Health care delivery to foster children: a study.

The health of 900 children in foster care was assessed through a review of 257 medical charts and an examination of a subsample of 35 children. Although major handicapping conditions among the children were well cared for, prevention, the care of minor conditions and emotional problems, and overall coordination of care were found to be lacking. On the basis of these findings, the authors propose how health care delivery to foster children can be improved.