STrain Analysis and Mapping of the Plantar Surface (STAMPS): A novel technique of plantar load analysis during gait

Diabetic foot ulceration is driven by peripheral neuropathy, resulting in abnormal foot biomechanics and elevated plantar load. Plantar load comprises normal pressure and tangential shear stress. Currently, there are no in-shoe devices measuring both components of plantar load. The STAMPS (STrain Analysis and Mapping of the Plantar Surface) system was developed to address this and utilises digital image correlation (DIC) to determine the strain sustained by a plastically deformable insole, providing an assessment of plantar load at the foot-surface interface during gait. STAMPS was developed as a multi-layered insole, comprising a deformable mid-layer, onto which a stochastic speckle pattern film is applied. A custom-built imaging platform is used to obtain high resolution pre- and post-walking images. Images are imported into commercially available DIC software (GOM Correlate, 2020) to obtain pointwise strain data. The strain and displacement data are exported and post-processed with custom analysis routines (MATLAB, Mathworks Inc.), to obtain the resultant global and regional peak strain (SMAG), antero-posterior strain (SAP) and medio-lateral strain (SML). To validate the core technique an experimental test process used a Universal Mechanical Tester (UMT) system (UMT TriboLab, Bruker) to apply controlled vertical and tangential load regimes to the proposed multi-layer insole. A pilot study was then conducted to assess the efficacy of using the STAMPS system to measure in-shoe plantar strain in three healthy participants. Each participant walked 10 steps on the STAMPS insole using a standardised shoe. They also walked 10 m in the same shoe using a plantar pressure measurement insole (Novel Pedar®) to record peak plantar pressure (PPP) as a gold-standard comparator. The results of the experimental validation tests show that with increased normal force, at a constant shear distance, SMAG increased in a linear fashion. Furthermore, they showed that with increased shear distance, at a constant force, SMAG increased. The results of the pilot study found participant 1 demonstrated greatest SMAG in the region toes 3–5 (15.31%). The highest mean SMAG for participant 2 was at the hallux (29.31%). Participant 3 exhibited highest strain in the regions of the first and second metatarsal heads (58.85% and 41.62% respectively). Increased PPP was strongly associated with increased SMAG with a Spearman’s correlation coefficient 0.673 (p < 0.0001). This study has demonstrated the efficacy of a novel method to assess plantar load across the plantar surface of the foot. Experimental testing validated the sensitivity of the method to both normal pressure and tangential shear stress. This technique was successfully incorporated into the STAMPS insole to reliably measure and quantify the cumulative degree of strain sustained by a plastically deformable insole during a period of gait, which can be used to infer plantar loading patterns. Future work will explore how these measures relate to different pathologies, such as regions at risk of diabetic foot ulceration

A RAC-GEF network critical for early intestinal tumourigenesis.

RAC1 activity is critical for intestinal homeostasis, and is required for hyperproliferation driven by loss of the tumour suppressor gene Apc in the murine intestine. To avoid the impact of direct targeting upon homeostasis, we reasoned that indirect targeting of RAC1 via RAC-GEFs might be effective. Transcriptional profiling of Apc deficient intestinal tissue identified Vav3 and Tiam1 as key targets. Deletion of these indicated that while TIAM1 deficiency could suppress Apc-driven hyperproliferation, it had no impact upon tumourigenesis, while VAV3 deficiency had no effect. Intriguingly, deletion of either gene resulted in upregulation of Vav2, with subsequent targeting of all three (Vav2-/- Vav3-/- Tiam1-/-), profoundly suppressing hyperproliferation, tumourigenesis and RAC1 activity, without impacting normal homeostasis. Critically, the observed RAC-GEF dependency was negated by oncogenic KRAS mutation. Together, these data demonstrate that while targeting RAC-GEF molecules may have therapeutic impact at early stages, this benefit may be lost in late stage disease

Tracking of autologous adipose tissue-derived mesenchymal stromal cells with in vivo magnetic resonance imaging and histology after intralesional treatment of artificial equine tendon lesions - a pilot study

Background: Adipose tissue-derived mesenchymal stromal cells (AT-MSCs) are frequently used to treat equine tendinopathies. Up to now, knowledge about the fate of autologous AT-MSCs after intralesional injection into equine superficial digital flexor tendons (SDFTs) is very limited. The purpose of this study was to monitor the presence of intralesionally injected autologous AT-MSCs labelled with superparamagnetic iron oxide (SPIO) nanoparticles and green fluorescent protein (GFP) over a staggered period of 3 to 9 weeks with standing magnetic resonance imaging (MRI) and histology. Methods: Four adult warmblood horses received a unilateral injection of 10 × 106 autologous AT-MSCs into surgically created front-limb SDFT lesions. Administered AT-MSCs expressed lentivirally transduced reporter genes for GFP and were co-labelled with SPIO particles in three horses. The presence of AT-MSCs in SDFTs was evaluated by repeated examinations with standing low-field MRI in two horses and post-mortem in all horses with Prussian blue staining, fluorescence microscopy and with immunofluorescence and immunohistochemistry using anti-GFP antibodies at 3, 5, 7 and 9 weeks after treatment. Results: AT-MSCs labelled with SPIO particles were detectable in treated SDFTs during each MRI in T2*- and T1-weighted sequences until the end of the observation period. Post-mortem examinations revealed that all treated tendons contained high numbers of SPIO- and GFP-labelled cells. Conclusions: Standing low-field MRI has the potential to track SPIO-labelled AT-MSCs successfully. Histology, fluorescence microscopy, immunofluorescence and immunohistochemistry are efficient tools to detect labelled AT-MSCs after intralesional injection into surgically created equine SDFT lesions. Intralesional injection of 10 × 106 AT-MSCs leads to the presence of high numbers of AT-MSCs in and around surgically created tendon lesions for up to 9 weeks. Integration of injected AT-MSCs into healing tendon tissue is an essential pathway after intralesional administration. Injection techniques have to be chosen deliberately to avoid reflux of the cell substrate injected. In vivo low-field MRI may be used as a non-invasive tool to monitor homing and engraftment of AT-MSCs in horses with tendinopathy of the SDFT

Oncogenic BRAF, unrestrained by TGFβ-receptor signalling, drives right-sided colonic tumorigenesis.

Right-sided (proximal) colorectal cancer (CRC) has a poor prognosis and a distinct mutational profile, characterized by oncogenic BRAF mutations and aberrations in mismatch repair and TGFβ signalling. Here, we describe a mouse model of right-sided colon cancer driven by oncogenic BRAF and loss of epithelial TGFβ-receptor signalling. The proximal colonic tumours that develop in this model exhibit a foetal-like progenitor phenotype (Ly6a/Sca1+) and, importantly, lack expression of Lgr5 and its associated intestinal stem cell signature. These features are recapitulated in human BRAF-mutant, right-sided CRCs and represent fundamental differences between left- and right-sided disease. Microbial-driven inflammation supports the initiation and progression of these tumours with foetal-like characteristics, consistent with their predilection for the microbe-rich right colon and their antibiotic sensitivity. While MAPK-pathway activating mutations drive this foetal-like signature via ERK-dependent activation of the transcriptional coactivator YAP, the same foetal-like transcriptional programs are also initiated by inflammation in a MAPK-independent manner. Importantly, in both contexts, epithelial TGFβ-receptor signalling is instrumental in suppressing the tumorigenic potential of these foetal-like progenitor cells

Failure to Detect the Novel Retrovirus XMRV in Chronic Fatigue Syndrome

BACKGROUND:In October 2009 it was reported that 68 of 101 patients with chronic fatigue syndrome (CFS) in the US were infected with a novel gamma retrovirus, xenotropic murine leukaemia virus-related virus (XMRV), a virus previously linked to prostate cancer. This finding, if confirmed, would have a profound effect on the understanding and treatment of an incapacitating disease affecting millions worldwide. We have investigated CFS sufferers in the UK to determine if they are carriers of XMRV. METHODOLOGY:Patients in our CFS cohort had undergone medical screening to exclude detectable organic illness and met the CDC criteria for CFS. DNA extracted from blood samples of 186 CFS patients were screened for XMRV provirus and for the closely related murine leukaemia virus by nested PCR using specific oligonucleotide primers. To control for the integrity of the DNA, the cellular beta-globin gene was amplified. Negative controls (water) and a positive control (XMRV infectious molecular clone DNA) were included. While the beta-globin gene was amplified in all 186 samples, neither XMRV nor MLV sequences were detected. CONCLUSION:XMRV or MLV sequences were not amplified from DNA originating from CFS patients in the UK. Although we found no evidence that XMRV is associated with CFS in the UK, this may be a result of population differences between North America and Europe regarding the general prevalence of XMRV infection, and might also explain the fact that two US groups found XMRV in prostate cancer tissue, while two European studies did not

The Myosin Va Head Domain Binds to the Neurofilament-L Rod and Modulates Endoplasmic Reticulum (ER) Content and Distribution within Axons

The neurofilament light subunit (NF-L) binds to myosin Va (Myo Va) in neurons but the sites of interaction and functional significance are not clear. We show by deletion analysis that motor domain of Myo Va binds to the NF-L rod domain that forms the NF backbone. Loss of NF-L and Myo Va binding from axons significantly reduces the axonal content of ER, and redistributes ER to the periphery of axon. Our data are consistent with a novel function for NFs as a scaffold in axons for maintaining the content and proper distribution of vesicular organelles, mediated in part by Myo Va. Based on observations that the Myo Va motor domain binds to intermediate filament (IF) proteins of several classes, Myo Va interactions with IFs may serve similar roles in organizing organelle topography in different cell types

Effects of Elevated Temperature and Carbon Dioxide on the Growth and Survival of Larvae and Juveniles of Three Species of Northwest Atlantic Bivalves

Rising CO2 concentrations and water temperatures this century are likely to have transformative effects on many coastal marine organisms. Here, we compared the responses of two life history stages (larval, juvenile) of three species of calcifying bivalves (Mercenaria mercenaria, Crassostrea virginica, and Argopecten irradians) to temperatures (24 and 28°C) and CO2 concentrations (∼250, 390, and 750 ppm) representative of past, present, and future summer conditions in temperate estuaries. Results demonstrated that increases in temperature and CO2 each significantly depressed survival, development, growth, and lipid synthesis of M. mercenaria and A. irradians larvae and that the effects were additive. Juvenile M. mercenaria and A. irradians were negatively impacted by higher temperatures while C. virginica juveniles were not. C. virginica and A. irradians juveniles were negatively affected by higher CO2 concentrations, while M. mercenaria was not. Larvae were substantially more vulnerable to elevated CO2 than juvenile stages. These findings suggest that current and future increases in temperature and CO2 are likely to have negative consequences for coastal bivalve populations

A hierarchy of effective teaching and learning to acquire competence in evidenced-based medicine

BACKGROUND: A variety of methods exists for teaching and learning evidence-based medicine (EBM). However, there is much debate about the effectiveness of various EBM teaching and learning activities, resulting in a lack of consensus as to what methods constitute the best educational practice. There is a need for a clear hierarchy of educational activities to effectively impart and acquire competence in EBM skills. This paper develops such a hierarchy based on current empirical and theoretical evidence. DISCUSSION: EBM requires that health care decisions be based on the best available valid and relevant evidence. To achieve this, teachers delivering EBM curricula need to inculcate amongst learners the skills to gain, assess, apply, integrate and communicate new knowledge in clinical decision-making. Empirical and theoretical evidence suggests that there is a hierarchy of teaching and learning activities in terms of their educational effectiveness: Level 1, interactive and clinically integrated activities; Level 2(a), interactive but classroom based activities; Level 2(b), didactic but clinically integrated activities; and Level 3, didactic, classroom or standalone teaching. SUMMARY: All health care professionals need to understand and implement the principles of EBM to improve care of their patients. Interactive and clinically integrated teaching and learning activities provide the basis for the best educational practice in this field

Preliminary outcomes of a paediatric highly active antiretroviral therapy cohort from KwaZulu-Natal, South Africa

BACKGROUND: Few studies address the use of paediatric highly active antiretroviral therapy (HAART) in Africa. METHODS: We performed a retrospective cohort study to investigate preliminary outcomes of all children eligible for HAART at Sinikithemba HIV/AIDS clinic in KwaZulu-Natal, South Africa. Immunologic, virologic, clinical, mortality, primary caregiver, and psychosocial variables were collected and analyzed. RESULTS: From August 31, 2003 until October 31, 2005, 151 children initiated HAART. The median age at HAART initiation was 5.7 years (range 0.3–15.4). Median follow-up time of the cohort after HAART initiation was 8 months (IQR 3.5–13.5). The median change in CD4% from baseline (p < 0.001) was 10.2 (IQR 5.0–13.8) at 6 months (n = 90), and 16.2 (IQR 9.6–20.3) at 12 months (n = 59). Viral loads (VLs) were available for 100 children at 6 months of which 84% had HIV-1 RNA levels ≤ 50 copies/mL. At 12 months, 80.3% (n = 61) had undetectable VLs. Sixty-five out of 88 children (73.8%) reported a significant increase (p < 0.001) in weight after the first month. Eighty-nine percent of the cohort (n = 132) reported ≤ 2 missed doses during any given treatment month (> 95%adherence). Seventeen patients (11.3%) had a regimen change; two (1.3%) were due to antiretroviral toxicity. The Kaplan-Meier one year survival estimate was 90.9% (95%confidence interval (CI) 84.8–94.6). Thirteen children died during follow-up (8.6%), one changed service provider, and no children were lost to follow-up. All 13 deaths occurred in children with advanced HIV disease within 5 months of treatment initiation. In multivariate analysis of baseline variables against mortality using Cox proportional-hazards model, chronic gastroenteritis was associated with death [hazard ratio (HR), 12.34; 95%CI, 1.27–119.71) and an HIV-positive primary caregiver was found to be protective against mortality [HR, 0.12; 95%CI, 0.02–0.88). Age, orphanhood, baseline CD4%, and hemoglobin were not predicators of mortality in our cohort. Fifty-two percent of the cohort had at least one HIV-positive primary caregiver, and 38.4% had at least one primary caregiver also on HAART at Sinikithemba clinic. CONCLUSION: This report suggests that paediatric HAART can be effective despite the challenges of a resource-limited setting

Abundance of Early Functional HIV-Specific CD8+ T Cells Does Not Predict AIDS-Free Survival Time

Background T-cell immunity is thought to play an important role in controlling HIV infection, and is a main target for HIV vaccine development. HIV-specific central memory CD8+ and CD4+ T cells producing IFNγ and IL-2 have been associated with control of viremia and are therefore hypothesized to be truly protective and determine subsequent clinical outcome. However, the cause-effect relationship between HIV-specific cellular immunity and disease progression is unknown. We investigated in a large prospective cohort study involving 96 individuals of the Amsterdam Cohort Studies with a known date of seroconversion whether the presence of cytokine-producing HIV-specific CD8+ T cells early in infection was associated with AIDS-free survival time. Methods and Findings The number and percentage of IFNγ and IL-2 producing CD8+ T cells was measured after in vitro stimulation with an overlapping Gag-peptide pool in T cells sampled approximately one year after seroconversion. Kaplan-Meier survival analysis and Cox proportional hazard models showed that frequencies of cytokine-producing Gag-specific CD8+ T cells (IFNγ, IL-2 or both) shortly after seroconversion were neither associated with time to AIDS nor with the rate of CD4+ T-cell decline. Conclusions These data show that high numbers of functional HIV-specific CD8+ T cells can be found early in HIV infection, irrespective of subsequent clinical outcome. The fact that both progressors and long-term non-progressors have abundant T cell immunity of the specificity associated with low viral load shortly after seroconversion suggests that the more rapid loss of T cell immunity observed in progressors may be a consequence rather than a cause of disease progression