Piscine (Sparus aurata) transthyretin cDNA cloning and characterization

Transthyretin (TTR) is one of the three plasma proteins that participate in the transport of thyroid hormones, thyroxine (T4) and triiodothyronine (T3); it is also involved in the carriage of retinol through the mediation of retinol-binding protein.1 The liver and choroid plexus are the major sites of TTR synthesis in mammals, birds, and diprotodont marsupials; in reptiles, TTR is only synthesized in the choroid plexus. 2 cDNA coding for TTR has been cloned from several mammalian, reptilian, and avian species and is highly conserved especially in the domains involved in binding to thyroid hormones.3 TTR expression has never been reported in fish, in which albumin is considered to be the main carrier for T3 and T4.2 We report the cloning of a TTR cDNA from sea bream (Sparus aurata) and demonstrate the presence of TTR mRNA in the liver of this fish

Vascular endothelial growth factor-D over-expressing tumor cells induce differential effects on uterine vasculature in a mouse model of endometrial cancer

BACKGROUND: It has been hypothesised that increased VEGF-D expression may be an independent prognostic factor for endometrial cancer progression and lymph node metastasis; however, the mechanism by which VEGF-D may promote disease progression in women with endometrial cancer has not been investigated. Our aim was to describe the distribution of lymphatic vessels in mouse uterus and to examine the effect of VEGF-D over-expression on these vessels in a model of endometrial cancer. We hypothesised that VEGF-D over-expression would stimulate growth of new lymphatic vessels into the endometrium, thereby contributing to cancer progression. METHODS: We initially described the distribution of lymphatic vessels (Lyve-1, podoplanin, VEGFR-3) and VEGF-D expression in the mouse uterus during the estrous cycle, early pregnancy and in response to estradiol-17beta and progesterone using immunohistochemistry. We also examined the effects of VEGF-D over-expression on uterine vasculature by inoculating uterine horns in NOD SCID mice with control or VEGF-D-expressing 293EBNA tumor cells. RESULTS: Lymphatic vessels positive for the lymphatic endothelial cell markers Lyve-1, podoplanin and VEGFR-3 profiles were largely restricted to the connective tissue between the myometrial circular and longitudinal muscle layers; very few lymphatic vessel profiles were observed in the endometrium. VEGF-D immunostaining was present in all uterine compartments (epithelium, stroma, myometrium), although expression was generally low. VEGF-D immunoexpression was slightly but significantly higher in estrus relative to diestrus; and in estradiol-17beta treated mice relative to vehicle or progesterone treated mice. The presence of VEGF-D over-expressing tumor cells did not induce endometrial lymphangiogenesis, although changes were observed in existing vessel profiles. For myometrial lymphatic and endometrial blood vessels, the percentage of profiles containing proliferating endothelial cells, and the cross sectional area of vessel profiles were significantly increased in response to VEGF-D in comparison to control tumor cells. In contrast, no significant changes were noted in myometrial blood vessels. In addition, examples of invading cells or tumor emboli were observed in mice receiving VEGF-D expressing 293EBNA cells. CONCLUSIONS: These results illustrate that VEGF-D over-expression has differential effects on the uterine vasculature. These effects may facilitate VEGF-D's ability to promote endometrial cancer metastasis and disease progression

Targeting lymphangiogenesis to prevent tumour metastasis

Recent studies involving animal models of cancer and clinicopathological analyses of human tumours suggest that the growth of lymphatic vessels (lymphangiogenesis) in or nearby tumours is associated with the metastatic spread of cancer. The best validated molecular signalling system for tumour lymphangiogenesis involves the secreted proteins vascular endothelial growth factor-C (VEGF-C) and VEGF-D that induce growth of lymphatic vessels via activation of VEGF receptor-3 (VEGFR-3) localised on the surface of lymphatic endothelial cells. In this review, we discuss the evidence supporting a role for this signalling system in the spread of cancer and potential approaches for blocking this system to prevent tumour metastasis

Phage-Derived Fully Human Monoclonal Antibody Fragments to Human Vascular Endothelial Growth Factor-C Block Its Interaction with VEGF Receptor-2 and 3

Vascular endothelial growth factor C (VEGF-C) is a key mediator of lymphangiogenesis, acting via its receptors VEGF-R2 and VEGF-R3. High expression of VEGF-C in tumors correlates with increased lymphatic vessel density, lymphatic vessel invasion, sentinel lymph node metastasis and poor prognosis. Recently, we found that in a chemically induced skin carcinoma model, increased VEGF-C drainage from the tumor enhanced lymphangiogenesis in the sentinel lymph node and facilitated metastatic spread of cancer cells via the lymphatics. Hence, interference with the VEGF-C/VEGF-R3 axis holds promise to block metastatic spread, as recently shown by use of a neutralizing anti-VEGF-R3 antibody and a soluble VEGF-R3 (VEGF-C/D trap). By antibody phage-display, we have developed a human monoclonal antibody fragment (single-chain Fragment variable, scFv) that binds with high specificity and affinity to the fully processed mature form of human VEGF-C. The scFv binds to an epitope on VEGF-C that is important for receptor binding, since binding of the scFv to VEGF-C dose-dependently inhibits the binding of VEGF-C to VEGF-R2 and VEGF-R3 as shown by BIAcore and ELISA analyses. Interestingly, the variable heavy domain (VH) of the anti-VEGF-C scFv, which contains a mutation typical for camelid heavy chain-only antibodies, is sufficient for binding VEGF-C. This reduced the size of the potentially VEGF-C-blocking antibody fragment to only 14.6 kDa. Anti-VEGF-C VH-based immunoproteins hold promise to block the lymphangiogenic activity of VEGF-C, which would present a significant advance in inhibiting lymphatic-based metastatic spread of certain cancer types

The Pain in Dystonia Scale (PIDS)-Development and Validation in Cervical Dystonia.

BACKGROUND: A better understanding of pain in adult-onset idiopathic dystonia (AOID) is needed to implement effective therapeutic strategies. OBJECTIVE: To develop a new rating instrument for pain in AOID and validate it in cervical dystonia (CD). METHODS: Development and validation of the Pain in Dystonia Scale (PIDS) comprised three phases. In phase 1, international experts and participants with AOID generated and evaluated the preliminary items for content validity. In phase 2, the PIDS was drafted and revised by the experts, followed by cognitive interviews to ensure self-administration suitability. In phase 3, the PIDS psychometric properties were assessed in 85 participants with CD and retested in 40 participants. RESULTS: The final version of PIDS evaluates pain severity (by body-part), functional impact, and external modulating factors. Test-retest reliability showed a high-correlation coefficient for the total score (0.9, P < 0.001), and intraclass correlation coefficients were 0.7 or higher for all items in all body-parts subscores. The overall PIDS severity score showed high internal consistency (Cronbach's α, 0.9). Convergent validity analysis revealed a strong correlation between the PIDS severity score and the Toronto Western Spasmodic Torticollis Rating Scale pain subscale (0.8, P < 0.001) and the Brief Pain Inventory-short form items related to pain at time of the assessment (0.7, P < 0.001) and impact of pain on daily functioning (0.7, P < 0.001). CONCLUSION: The PIDS is the first specific questionnaire developed to evaluate pain in all patients with AOID, here, demonstrating high-level psychometric properties in people with CD. Future work will validate PIDS in other forms of AOID. © 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society

Mechanism of IL-12 mediated alterations in tumour blood vessel morphology: analysis using whole-tissue mounts

Angiogenesis is a multistep process that is limited and carefully regulated in normal adult tissue, but in tumours this regulation is disrupted and the process remains ‘switched on’ (Hanahan and Folkman, 1996). Ample experimental data support the fact that tumour growth requires access to blood vessels and subsequent expansion of host vessels to provide nutrients for the growing tumour mass (Folkman, 1995a). Furthermore, many studies in a variety of tumour types have reported a correlation between the extent of tumour vasculature and poor prognosis or increased metastases (Weidner et al, 1991; Folkman, 1995b; Weidner and Folkman, 1996). Thus, accurate assessment of the vasculature of tumours could provide valuable information regarding treatment outcomes and the likelihood of metastatic spread to other sites. Angiogenesis can be regulated by a variety of factors. Several cytokines produced by immune cells also have been shown to affect the process of angiogenesis. One of the most noteworthy is interleukin (IL)-12, which is produced by antigen presenting cells (APC), such as macrophages and dendritic cells (DC) in response to bacterial stimuli or other inflammatory cytokines. Thus, IL-12 plays an important role in both the innate and adaptive immune responses (Trinchieri, 1998). Owing to its central role in stimulating immunity, it has been examined for possible therapeutic effects in the treatment of tumours. In addition to its effects on the immune system, IL-12 has also been shown to inhibit angiogenesis (Voest et al, 1995; Sgadari et al, 1996). Despite studies in both experimental models and in patients (reviewed in Trinchieri and Scott, 1999), and clear demonstrations of therapeutic efficacy, relatively little is known about how it alters vessel formation within tumours. In part, this is due to the difficulty in assessing the three-dimensional structure of vessels and other cellular components within the tumour. Assessment of tumour vessels is generally based on immunohistochemistry of tumour sections. Although use of this technique has led to a great deal of important information, these procedures are extremely time consuming and provide only a limited two-dimensional view of the vessels. This makes it very difficult to visualise the structure of the microvasculature and identify differences among different tumour types or changes following treatment regimens. To more easily and accurately visualise vessels within tumours, we developed a whole-tissue mount technique that provides a three-dimensional view of the tumour vasculature relative to other components of the tumour tissue. This technique was first validated by studying vessels from transgenic mice that express green fluorescent protein (GFP) (Wu et al, 2000), and then used to investigate the mechanism by which IL-12 influences the vessel architecture within B16 tumours

The Prevalence of Campylobacter amongst a Free-Range Broiler Breeder Flock Was Primarily Affected by Flock Age

Campylobacter successfully colonizes broiler chickens, but little is known about the longer term natural history of colonization, since most flocks are slaughtered at an immature age. In this study, the prevalence and genetic diversity of Campylobacter colonizing a single free-range broiler breeder flock was investigated over the course of a year. The age of the flock was the most important factor in determining both the prevalence and diversity of Campylobacter over time. There was no correlation with season, temperature, the amount of rain and sunshine, or the dynamics of colonization amongst geographically and temporally matched broiler flocks. The higher prevalence rates coincided with the age at which broiler chickens are typically slaughtered, but then in the absence of bio-security or other intervention methods, and despite changes in flock management, the prevalence fell to significantly lower levels for the remainder of the study. The genetic diversity of Campylobacter increased as the flock aged, implying that genotypes were accumulated within the flock and may persist for a long time. A better understanding of the ecology of Campylobacter within commercial chicken flocks will allow the design of more effective farm-based interventions