Response-guided neoadjuvant sacituzumab govitecan for localized triple-negative breast cancer: results from the NeoSTAR trial.

BACKGROUND: Sacituzumab govitecan (SG), a novel antibody-drug conjugate (ADC) targeting TROP2, is approved for pre-treated metastatic triple-negative breast cancer (mTNBC). We conducted an investigator-initiated clinical trial evaluating neoadjuvant (NA) SG (NCT04230109), and report primary results. PATIENTS AND METHODS: Participants with early-stage TNBC received NA SG for four cycles. The primary objective was to assess pathological complete response (pCR) rate in breast and lymph nodes (ypT0/isN0) to SG. Secondary objectives included overall response rate (ORR), safety, event-free survival (EFS), and predictive biomarkers. A response-guided approach was utilized, and subsequent systemic therapy decisions were at the discretion of the treating physician. RESULTS: From July 2020 to August 2021, 50 participants were enrolled (median age = 48.5 years; 13 clinical stage I disease, 26 stage II, 11 stage III). Forty-nine (98%) completed four cycles of SG. Overall, the pCR rate with SG alone was 30% [n = 15, 95% confidence interval (CI) 18% to 45%]. The ORR per RECIST V1.1 after SG alone was 64% (n = 32/50, 95% CI 77% to 98%). Higher Ki-67 and tumor-infiltrating lymphocytes (TILs) were predictive of pCR to SG (P = 0.007 for Ki-67 and 0.002 for TILs), while baseline TROP2 expression was not (P = 0.440). Common adverse events were nausea (82%), fatigue (76%), alopecia (76%), neutropenia (44%), and rash (48%). With a median follow-up time of 18.9 months (95% CI 16.3-21.9 months), the 2-year EFS for all participants was 95%. Among participants with a pCR with SG (n = 15), the 2-year EFS was 100%. CONCLUSIONS: In the first NA trial with an ADC in localized TNBC, SG demonstrated single-agent efficacy and feasibility of response-guided escalation/de-escalation. Further research on optimal duration of SG as well as NA combination strategies, including immunotherapy, are needed.

Late diagnosis of tuberous sclerosis: a case report.

Tuberous sclerosis complex (TSC) is an autosomal dominant genetic disorder characterized by the formation of hamartomas in organ systems such as the brain, skin, kidneys and lungs. Patients with TSC are usually diagnosed early in life. However, in some cases, the diagnosis is delayed until adulthood because various manifestations occur at various times throughout an individual's life. In this regard, we present the case of a female patient diagnosed at the beginning of the seventh decade of life. The patient had a history of seizures and showed clinical findings on the skin (facial angiofibromas, ungual fibromas, 'Confetti-like' skin lesions, shagreen patch), brain (cortical tubers), heart (cardiac rhabdomyomas), kidneys (angiomyolipomas) and a positive genetic test for mutations in TSC2, fulfilling the diagnostic criteria. We compared the differences between manifestations in patients diagnosed during childhood and adulthood. Knowledge of the clinical spectrum of TSC allows early identification.

Implementing surgical mentorship in a resource-constrained context: a mixed methods assessment of the experiences of mentees, mentors, and leaders, and lessons learned.

BACKGROUND: A well-qualified workforce is critical to effective functioning of health systems and populations; however, skill gaps present a challenge in low-resource settings. While an emerging body of evidence suggests that mentorship can improve quality, access, and systems in African health settings by building the capacity of health providers, less is known about its implementation in surgery. We studied a novel surgical mentorship intervention as part of a safe surgery intervention (Safe Surgery 2020) in five rural Ethiopian facilities to understand factors affecting implementation of surgical mentorship in resource-constrained settings. METHODS: We designed a convergent mixed-methods study to understand the experiences of mentees, mentors, hospital leaders, and external stakeholders with the mentorship intervention. Quantitative data was collected through a survey (n = 25) and qualitative data through in-depth interviews (n = 26) in 2018 to gather information on (1) intervention characteristics including areas of mentorship, mentee-mentor relationships, and mentor characteristics, (2) organizational context including facilitators and barriers to implementation, (3) perceived impact, and (4) respondent characteristics. We analyzed the quantitative and qualitative data using frequency analysis and the constant comparison method, respectively; we integrated findings to identify themes. RESULTS: All mentees (100%) experienced the intervention as positive. Participants perceived impact as: safer and more frequent surgical procedures, collegial bonds between mentees and mentors, empowerment among mentees, and a culture of continuous learning. Over 70% of all mentees reported their confidence and job satisfaction increased. Supportive intervention characteristics included a systems focus, psychologically safe mentee-mentor relationships, and mentor characteristics including generosity with time and knowledge, understanding of local context, and interpersonal skills. Supportive organizational context included a receptive implementation climate. Intervention challenges included insufficient clinical training, inadequate mentor support, and inadequate dose. Organizational context challenges included resource constraints and a lack of common understanding of the intervention. CONCLUSION: We offer lessons for intervention designers, policy makers, and practitioners about optimizing surgical mentorship interventions in resource-constrained settings. We attribute the intervention's success to its holistic approach, a receptive climate, and effective mentee-mentor relationships. These qualities, along with policy support and adapting the intervention through user feedback are important for successful implementation.

Congruencia entre modelos dentales impresos 3D y el archivo digital de origen: estudio in vitro comparativo de 5 impresoras 3D / Congruence between 3D printed dental models and the original digital file: comparative in vitro study of 5 3D printers

Este trabajo tuvo como objetivo conocer la fiabilidad de la impresora 3D (i3D) aditiva por Matriz de Proceso Digital de Luz (MDLP) Hellbot modelo Apolo®, a través de verificar la congruencia dimensional entre las mallas de modelos impresos (MMi) y su correspondiente archivo digital de origen (MMo), obtenido del software de planificación ortodontica Orchestrate 3D® (O3D). Para determinar su uso en odontología y sus posibilidades clínicas, fue comparada entre cinco i3D de manufactura aditiva, dos DLP, dos por estereolitografía (SLA) y una por Depósito de Material Fundido (FDM). La elección de las cinco i3D se fundamentó en su valor de mercado, intentando abarcar la mayor diversidad argentina disponible. Veinte modelos fueron impresos con cada i3D y escaneados con Escáner Intraoral (IOS) Carestream modelo 3600® (Cs3600). Las 120 MMi fueron importadas dentro del programa de ingeniería inversa Geomagic® Control X® (Cx) para su análisis 3D, consistiendo en la superposición de MMo con cada una de las MMi. Luego, una evaluación cualitativa de la desviación entre la MMi y MMo fue realizada. Un análisis estadístico cuidadoso fue realizado obteniendo como resultado comparaciones en 3d y 2d. Las coincidencias metrológicas en la superposición tridimensional permitieron un análisis exhaustivo y fácilmente reconocible a través de mapas colorimétricos. En el análisis bidimensional se plantearon planos referenciados dentariamente desde la MMo, para hacer coincidir las mediciones desde el mismo punto de partida dentaria. Los resultados fueron satisfactorios y muy alentadores. Las probabilidades de obtener rangos de variabilidad equivalentes a +/- 50µm fueron de un 40,35 % y de +/- 100µm un 71,04 %. Por lo tanto, te- niendo en cuenta las exigencias de congruencia dimensional clínicas de precisión y exactitud a las cuales es sometida nuestra profesión odontológica, se evitan problemas clínicos arrastrados por los errores dimensionales en la manufactura (Cam) (AU)

The objective of this study was to determine the reliability of the Hellbot Apollo® model additive 3D printer (i3D) by Matrix Digital Light Processing (MDLP) by verifying the dimensional congruence between the printed model meshes (MMi) and their corresponding digital source file (MMo), obtained from the Orchestrate 3D® (O3D) orthodontic planning software. A comparison was made between five i3D of additive manufacturing, two DLP, two by stereolithography (SLA), and one by Fused Material Deposition (FDM), to determine its use in dentistry and its clinical possibilities. The choice of the five i3D was based on their market value, trying to cover most of the Argentinean diversity available. Twenty models were printed with each i3D and scanned with Carestream Intraoral Scanner (IOS) model 3600® (Cs3600). The 120 MMi were imported into the reverse engineering program Geomagic® Control X® (Cx) for 3D analysis, consisting of overlaying MMo with each MMi. Then, a qualitative evaluation of the deviation between MMi and MMo. Also, a careful statistical analysis was performed, resulting in 3d and 2d comparisons. Metrological coincidences in three-dimensional overlay allowed a comprehensive and easily recognizable analysis through colorimetric maps. In the two-dimensional analysis, dentally referenced planes were proposed from the MMo, to match the measurements from the same dental starting point. The results were satisfactory and very encouraging. The probabilities of obtaining ranges of variability equivalent to +/- 50µm were 40.35 % and +/- 100µm 71.04 %. Therefore, considering the demands of clinical dimensional congruence, precision, and accuracy to which our dental profession it is subjected, clinical problems caused by dimensional errors in manufacturing (Cam) are avoided (AU)

Mental health improvements after elective spine surgery: a Canadian Spine Outcome Research Network (CSORN) study.

BACKGROUND CONTEXT: Spine patients have a higher rate of depression then the general population which may be caused in part by levels of pain and disability from their spinal disease. PURPOSE: Determination whether improvements in health-related quality of life (HRQOL) resulting from successful spine surgery leads to improvements in mental health. STUDY DESIGN/SETTING: The Canadian Spine Outcome Research Network prospective surgical outcome registry. OUTCOME MEASURES: Change between preoperative and postoperative SF12 Mental Component Score (MCS). Secondary outcomes include European Quality of Life (EuroQoL) Healthstate, SF-12 Physical Component Score (PCS), Oswestry Disability Index (ODI), Patient Health Questionaire-9 (PHQ9), and pain scales. METHODS: The Canadian Spine Outcome Research Network registry was queried for all patients receiving surgery for degenerative thoracolumbar spine disease. Exclusion criteria were trauma, tumor, infection, and previous spine surgery. SF12 Mental Component Scores (MCS) were compared between those with and without significant improvement in postoperative disability (ODI) and secondary measures. Multivariate analysis examined factors predictive of MCS improvement. RESULTS: Eighteen hospitals contributed 3222 eligible patients. Worse ODI, EuroQoL, PCS, back pain and leg pain correlated with worse MCS at all time points. Overall, patients had an improvement in MCS that occurred within 3 months of surgery and was still present 24 months after surgery. Patients exceeding Minimally Clinically Important Differences in ODI had the greatest improvements in MCS. Major depression prevalence decreased up to 48% following surgery, depending on spine diagnosis. CONCLUSIONS: Large scale, real world, registry data suggests that successful surgery for degenerative lumbar disease is associated with reduction in the prevalence of major depression regardless of the specific underlaying diagnosis. Worse baseline MCS was associated with worse baseline HRQOL and improved postoperatively with coincident improvement in disability, emphasizing that mental wellness is not a static state but may improve with well-planned spine surgery.

Epithelial tissue geometry directs emergence of bioelectric field and pattern of proliferation.

Patterns of proliferation are templated by both gradients of mechanical stress as well as by gradients in membrane voltage (Vm), which is defined as the electric potential difference between the cytoplasm and the extracellular medium. Either gradient could regulate the emergence of the other, or they could arise independently and synergistically affect proliferation within a tissue. Here, we examined the relationship between endogenous patterns of mechanical stress and the generation of bioelectric gradients in mammary epithelial tissues. We observed that the mechanical stress gradients in the tissues presaged gradients in both proliferation and depolarization, consistent with previous reports correlating depolarization with proliferation. Furthermore, disrupting the Vm gradient blocked the emergence of patterned proliferation. We found that the bioelectric gradient formed downstream of mechanical stresses within the tissues and depended on connexin-43 (Cx43) hemichannels, which opened preferentially in cells located in regions of high mechanical stress. Activation of Cx43 hemichannels was necessary for nuclear localization of Yap/Taz and induction of proliferation. Together, these results suggest that mechanotransduction triggers the formation of bioelectric gradients across a tissue, which are further translated into transcriptional changes that template patterns of growth.

PARATHYROMATOSIS: A RARE CASE OF RECURRENT HYPERPARATHYROIDISM LOCALIZED BY FOUR-DIMENSIONAL COMPUTED TOMOGRAPHY.

OBJECTIVE: To present a rare case of parathyromatosis. METHODS: We present the clinical, laboratory, and imaging findings, along with a review of the literature. RESULTS: A 33-year-old man with a history of right upper parathyroid adenoma removal 5 years prior due to hyperparathyroidism was admitted for severe hypercalcemia (15.6 mg/dL; normal, 8.5 to 10.5 mg/dL) with elevated plasma parathyroid hormone (PTH) (882 pg/mL; normal, 15 to 65 pg/mL). Ultrasound, computed tomography (CT), sestamibi, and positron emission tomography scans were unremarkable; however, a four-dimensional CT (4DCT) of the neck showed an area of increased signal enhancement and hypervascularity without discrete nodule in the posterior right thyroid region. The patient underwent parathyroid surgical exploration with right hemithyroidectomy and compartment neck dissection to remove the affected tissue. PTH levels dropped to 208 pg/mL postoperatively; calcium decreased but remained elevated at 12.7 mg/dL. Pathology revealed the presence of several small nodular foci of atypical hyperplastic parathyroid tissue in the right thyroid and soft tissue in the left central neck compartment consistent with parathyromatosis. CONCLUSION: This case report represents the first-time use of 4DCT to localize parathyromatosis. Parathyromatosis is a rare but problematic cause of recurrent hyperparathyroidism. Ultrasound and 4DCT may represent the best imaging modalities for identification and perioperative management to remove all affected tissue without reseeding.

Dynamics of Tissue-Induced Alignment of Fibrous Extracellular Matrix.

Aligned fibers of extracellular matrix (ECM) affect the direction, efficiency, and persistence of migrating cells. To uncover the mechanisms by which multicellular tissues align their surrounding ECM before migration, we used an engineered three-dimensional culture model to investigate the dynamics of ECM alignment around tissues of defined geometry. Analysis of ECM alignment over time revealed that tissues rapidly reorganize their surrounding matrix, with a characteristic time that depends on the type of cell and the initial tissue geometry. We found that matrix metalloproteinase activity is not required for matrix alignment before cell migration. Instead, alignment is driven by Rho-mediated cytoskeletal contractility and accelerated by propagation of tension through intercellular adhesions. Our data suggest that multicellular tissues align their surrounding matrix by pulling collectively to exert strain, which is primarily a physical process. Consistently, the pattern of matrix alignment depends on tissue geometry and the resulting distribution of mechanical strain, with asymmetric tissues generating a higher degree of matrix alignment along their longest axes. The rapid ability of multicellular tissues to physically remodel their matrix enables their constituent cells to migrate efficiently along aligned fibers and to quickly change their direction according to other microenvironmental cues, which is important for both normal and disease processes.

Reported prevalence of gestational diabetes in Scotland: The relationship with obesity, age, socioeconomic status, smoking and macrosomia, and how many are we missing?

AIMS/INTRODUCTION: Gestational diabetes mellitus (GDM) is defined as 'carbohydrate intolerance of varying degrees of severity with onset or first recognition during pregnancy,' and is associated with increased fetal and maternal risks. The aims of the present study were to investigate the prevalence of GDM in Scotland over 32 years (1981-2012), and using the data from 2012, to assess how GDM related to maternal body mass index, maternal age, parity, smoking, Scottish Index of Multiple Deprivation, infant gender and macrosomia status. MATERIALS AND METHODS: GDM prevalence along with anthropometric, obstetric and demographic data were collected on a total of 1,891,097 women with a delivery episode between 1 January 1981 and 31 December 2012 using data extracted from the Scottish Morbidity Record 02. Univariate and multivariate logistic regression analysis was undertaken to investigate their association with GDM. RESULTS: A ninefold increase in GDM prevalence was observed from 1981 to 2012 (P < 0.001). GDM prevalence in 2012 was 1.9%. Maternal body mass index, age, parity status, Scottish index of multiple deprivation and fetal macrosomia were positively associated with GDM. Reported smoking status at booking was inversely associated with GDM. Multivariable analysis showed that fetal macrosomia was not associated with GDM status. CONCLUSIONS: The present study confirmed that the reporting of GDM is low in Scotland, and that GDM is associated with maternal body mass index, maternal age, multiparity and social deprivation. GDM was negatively associated with smoking and requires further investigation. The lack of association between GDM and macrosomia (following multivariate analysis) might reflect the screening processes undertaken in Scotland.

Chemoprotective effect of Vernonia amygdalina Del. (Astereacea) against 2-acetylaminofluorene-induced hepatotoxicity in rats.

Natural products possessing antioxidant properties play a very crucial role in ameliorating deleterious effects of reactive oxygen species. This study investigated the chemoprotective properties of methanolic extract of Vernonia amygdalina (MEVA) in an experimental model of hepatic oxidative damage induced by 2-acetylaminofluorene (2-AAF). Rats were divided into six groups. Groups 1 and 2 received saline and dimethyl sulfoxide, respectively, and served as controls. Group 3 received MEVA at a dose of 250 mg/kg, while groups 5 and 6 were pretreated for 14 days with MEVA at 250 mg/kg and 500 mg/kg doses before coadministration with 2-AAF at 100 mg/kg for another 7 days. 2-AAF was administered to group 4 for the last 7 days. Animals were killed 24 h after the last administration of 2-AAF. 2-AAF significantly (p < 0.05) induced marked hepatic damage as revealed by increased activities of serum enzymes such as alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, and γ-glutamyl transferase and bilirubin concentration. 2-AAF also elicited decrease in the activities of antioxidant enzymes such as superoxide dismutase, catalase, glutathione-S-transferase, and glutathione peroxidase, depletion of reduced glutathione, and increase in malondialdehyde levels. The activities of glucose-6-phosphatase and 5'-nucleotidase were also depleted. MEVA at 250 mg/kg and 500 mg/kg significantly (p < 0.05) ameliorated the oxidative damage, functional impairments, and histopathological changes associated with 2-AAF toxicity by reducing the activities of serum enzymes, upregulating the antioxidant defense enzymes and glutathione with decrease in malondialdehyde level. In this study, the revealed ameliorative and hepatoprotective effects of MEVA against 2-AAF-induced toxicity may be due to its antioxidant and free-radical scavenging activities, thus suggesting its usefulness as a possible chemoprophylactic agent.

Vascular Dilation, Tachycardia, and Increased Inotropy Occur Sequentially with Increasing Epinephrine Dose Rate, Plasma and Myocardial Concentrations, and cAMP.

BACKGROUND: While epinephrine infusion is widely used in critical care for inotropic support, there is no direct method to detect the onset and measure the magnitude of this response. We hypothesised that surrogate measurements, such as heart rate and vascular tone, may indicate if the plasma and tissue concentrations of epinephrine and cAMP are in a range sufficient to increase myocardial contractility. METHODS: Cardiovascular responses to epinephrine infusion (0.05-0.5 mcgkg(-1)min(-1)) were measured in rats using arterial and left ventricular catheters. Epinephrine and cAMP levels were measured using ELISA techniques. RESULTS: The lowest dose of epinephrine infusion (0.05 mcgkg(-1)min(-1)) did not raise plasma epinephrine levels and did not lead to cardiovascular response. Incremental increase in epinephrine infusion (0.1 mcgkg(-1)min(-1)) elevated plasma but not myocardial epinephrine levels, providing vascular, but not cardiac effects. Further increase in the infusion rate (0.2 mcgkg(-1)min(-1)) raised myocardial tissue epinephrine levels sufficient to increase heart rate but not contractility. Inotropic and lusitropic effects were significant at the infusion rate of 0.3 mcgkg(-1)min(-1). Correlation of plasma epinephrine to haemodynamic parameters suggest that as plasma concentration increases, systemic vascular resistance falls (EC50=47 pg/ml), then HR increases (ED50=168 pg/ml), followed by a rise in contractility and lusitropy (ED50=346 pg/ml and ED50=324 pg/ml accordingly). CONCLUSIONS: The dose response of epinephrine is distinct for vascular tone, HR and contractility. The need for higher doses to see cardiac effects is likely due to the threshold for drug accumulation in tissue. Successful inotropic support with epinephrine cannot be achieved unless the infusion is sufficient to raise the heart rate.

High concentrations of drug in target tissues following local controlled release are utilized for both drug distribution and biologic effect: an example with epicardial inotropic drug delivery.

Local drug delivery preferentially loads target tissues with a concentration gradient from the surface or point of release that tapers down to more distant sites. Drug that diffuses down this gradient must be in unbound form, but such drug can only elicit a biologic effect through receptor interactions. Drug excess loads tissues, increasing gradients and driving penetration, but with limited added biological response. We examined the hypothesis that local application reduces dramatically systemic circulating drug levels but leads to significantly higher tissue drug concentration than might be needed with systemic infusion in a rat model of local epicardial inotropic therapy. Epinephrine was infused systemically or released locally to the anterior wall of the heart using a novel polymeric platform that provides steady, sustained release over a range of precise doses. Epinephrine tissue concentration, upregulation of cAMP, and global left ventricular response were measured at equivalent doses and at doses equally effective in raising indices of contractility. The contractile stimulation by epinephrine was linked to drug tissue levels and commensurate cAMP upregulation for IV systemic infusion, but not with local epicardial delivery. Though cAMP was a powerful predictor of contractility with local application, tissue epinephrine levels were high and variable--only a small fraction of the deposited epinephrine was utilized in second messenger signaling and biologic effect. The remainder of deposited drug was likely used in diffusive transport and distribution. Systemic side effects were far more profound with IV infusion which, though it increased contractility, also induced tachycardia and loss of systemic vascular resistance, which were not seen with local application. Local epicardial inotropic delivery illustrates then a paradigm of how target tissues differentially handle and utilize drug compared to systemic infusion.

Involvement of hypoxia-inducible factor-1 in the inflammatory responses of human LAD2 mast cells and basophils.

We recently showed that hypoxia-inducible factor 1 (HIF-1) plays a crucial role in the pro-allergic functions of human basophils by transcriptional control of energy metabolism via glycolysis as well as directly triggering expression of the angiogenic cytokine vascular endothelium growth factor (VEGF). Here, we investigated HIF-1 involvement in controlling the synthesis of angiogenic and inflammatory cytokines from various human effector cells stimulated by IgE-dependent or innate immune triggers. Purified primary human basophils, LAD2 human mast cells and THP-1 human myeloid cells were used for investigations of FcεRI and Toll-like receptor (TLR) ligand-induced responses. In contrast to basophils, LAD2 mast cells expressed background levels of HIF-1α, which was largely independent of the effects of stem cell factor (SCF). Both mast cells and basophils expressed TLR2 and 4, albeit weakly compared to THP-1 cells. Cytokine production in mast cells following TLR ligand stimulation was markedly reduced by HIF-1α knockdown in LAD2 mast cells. In contrast, although HIF-1 is involved in IgE-mediated IL-4 secretion from basophils, it is not clearly induced by peptidoglycan (PGN). HIF-1α accumulation is critical for sustaining human allergic effector cell survival and function. This transcription complex facilitates generation of both pro-angiogenic and inflammatory cytokines in mast cells but has a differential role in basophil stimulation comparing IgE-dependent triggering with innate immune stimuli.

p53 Superfamily proteins in marine bivalve cancer and stress biology.

The human p53 tumour suppressor protein is inactivated in many cancers and is also a major player in apoptotic responses to cellular stress. The p53 protein and the two other members of this protein family (p63, p73) are encoded by distinct genes and their functions have been extensively documented for humans and some other vertebrates. The structure and relative expression levels for members of the p53 superfamily have also been reported for most major invertebrate taxa. The functions of homologous proteins have been investigated for only a few invertebrates (specifically, p53 in flies, nematodes and recently a sea anemone). These studies of classical model organisms all suggest that the gene family originally evolved to mediate apoptosis of damaged germ cells or to protect germ cells from genotoxic stress. Here, we have correlated data from a number of molluscan and other invertebrate sequencing projects to provide a framework for understanding p53 signalling pathways in marine bivalve cancer and stress biology. These data suggest that (a) the two identified p53 and p63/73-like proteins in soft shell clam (Mya arenaria), blue mussel (Mytilus edulis) and Northern European squid (Loligo forbesi) have identical core sequences and may be splice variants of a single gene, while some molluscs and most other invertebrates have two or more distinct genes expressing different p53 family members; (b) transcriptional activation domains (TADs) in bivalve p53 and p63/73-like protein sequences are 67-69% conserved with human p53, while those in ecdysozoan, cnidarian, placozoan and choanozoan eukaryotes are ≤33% conserved; (c) the Mdm2 binding site in the transcriptional activation domain is 100% conserved in all sequenced bivalve p53 proteins (e.g. Mya, Mytilus, Crassostrea and Spisula) but is not present in other non-deuterostome invertebrates; (d) an Mdm2 homologue has been cloned for Mytilus trossulus; (e) homologues for both human p53 upstream regulatory and transcriptional target genes exist in molluscan genomes (missing are ARF, CIP1 and BH3 only proteins) and (f) p53 is demonstrably involved in bivalve haemocyte and germinoma cancers. We usually do not know enough about the molecular biology of marine invertebrates to address molecular mechanisms that characterize particular diseases. Understanding the molecular basis of naturally occurring diseases in marine bivalves is a virtually unexplored aspect of toxicoproteomics and genomics and related drug discovery. Additionally, increases in coastal development and concomitant increases in aquatic pollutants have driven interest in developing models appropriate for evaluating potential hazardous compounds or conditions found in the aquatic environment. Data reviewed in this study are coupled with recent developments in our understanding the molecular biology of the marine bivalve p53 superfamily. Taken together, they suggest that both structurally and functionally, bivalve p53 family proteins are the most highly conserved members of this gene superfamily so far identified outside of higher vertebrates and invertebrate chordates. Marine bivalves provide some of the most relevant and best understood models currently available for experimental studies by biomedical and marine environmental researchers.

Effects of stem cell factor on hypoxia-inducible factor 1 alpha accumulation in human acute myeloid leukaemia and LAD2 mast cells.

Stem cell factor (SCF) is a hematopoietic growth factor that exerts its activity by signalling through the tyrosine kinase receptor known as Kit or CD117. SCF-Kit signalling is crucial for the survival, proliferation and differentiation of hematopoietic cells of myeloid lineage. Furthermore, since myeloid leukaemia cells express the Kit receptor, SCF may play an important role in myeloid leukaemia progression too. However, the mechanisms of this pathophysiological effect remain unclear. Recent evidence shows that SCF triggers accumulation of the inducible alpha subunit of hypoxia-inducible factor 1 (HIF-1) in hematopoietic cells--a transcription complex that plays a pivotal role in cellular adaptation to low oxygen availability. However, it is unknown how SCF impacts on HIF-1α accumulation in human myeloid leukaemia and mast cells. Here we show that SCF induces HIF-1α accumulation in THP-1 human myeloid leukaemia cells but not in LAD2 mast cells. We demonstrated that LAD2 cells have a more robust glutathione (GSH)-dependent antioxidative system compared to THP-1 cells and are therefore protected against the actions of ROS generated in an SCF-dependent manner. BSO-induced GSH depletion led to a significant decrease in HIF-1α prolyl hydroxylase (PHD) activity in THP-1 cells and to near attenuation of it in LAD2 cells. In THP-1 cells, SCF-induced HIF-1α accumulation is controlled via ERK, PI3 kinase/PKC-δ/mTOR-dependent and to a certain extent by redox-dependent mechanisms. These results demonstrate for the first time an important cross-talk of signalling pathways associated with HIF-1 activation--an important stage of the myeloid leukaemia cell life cycle.

Myeloid cell death associated with Toll-like receptor 7/8-mediated inflammatory response. Implication of ASK1, HIF-1 alpha, IL-1 beta and TNF-alpha.

Programmed cell death or apoptosis is an important part of the host innate immune defence, especially against ssRNA viruses (influenza virus, HIV-1, ebola virus, hepatitis C virus and many others). Viral ssRNA is recognised by endosomal Toll-like receptors 7 and 8 (TLR7/8) which induce further stages of immune defence against these pathogens. Some of the immune cells die because of inflammatory stress allowing for the selection of those cells which are resistant to stress-induced apoptosis and which are used in further stages of the host immune response. On the other hand, apoptosis could be used as an instrument to suppress the function of activated inflammatory cells. However, the mechanisms underlying death of the inflammatory cells associated with stress induced by ligands of TLR7/8 remain unclear. In this study we have found that programmed death of human myeloid cells from different cell lines associated with ligand-induced TLR7/8-mediated inflammatory stress depends on activation of apoptosis signal-regulating kinase 1 (ASK1). This enzyme is, however, not required for the production of pro-inflammatory cytokines - TNF-α and IL-1ß. We have found that released IL-1ß and TNF-α are involved in apoptosis of myeloid cells associated with TLR7/8-mediated inflammatory stress. The pro-apoptotic effect of released TNF-α in this case is much lower compared to that of IL-1ß.

Early development of forebrain gonadotrophin-releasing hormone (GnRH) neurones and the role of GnRH as an autocrine migration factor.

Normal migration of the gonadotrophin-releasing hormone (GnRH) neurones during early development, from the olfactory region to the hypothalamus, is crucial for reproductive development in all vertebrates. The establishment of the GnRH system includes tangential migration of GnRH perikarya as well as extension of GnRH fibres to various areas of the central nervous system (CNS). The exact spatio-temporal nature of this process, as well as the factors governing it, are not fully understood. We studied the development of the GnRH system and the effects of GnRH knockdown using a newly developed GnRH3:EGFP transgenic zebrafish line. We found that enhanced green fluorescent protein is specifically and robustly expressed in GnRH3 neurones and fibres. GnRH3 fibres in zebrafish began to extend as early as 26 h post-fertilisation and by 4-5 days post-fertilisation had developed into an extensive network reaching the optic tract, telencephalon, hypothalamus, midbrain tegmentum and hindbrain. GnRH3 fibres also innervated the retina and projected into the trunk via the spinal cord. GnRH3 perikarya were observed migrating along their own fibres from the olfactory region to the preoptic area (POA) via the terminal nerve ganglion and the ventral telencephalon. GnRH3 cells were also observed in the trigeminal ganglion. The establishment of the GnRH3 fibre network was disrupted by morpholino-modified antisense oligonucleotides directed against GnRH3 causing abnormal fibre development and pathfinding, as well as anomalous GnRH3 perikarya localisation. These findings support the hypothesis that GnRH3 neurones migrate from the olfactory region to the POA and caudal hypothalamus. Novel data regarding the early development of the GnRH3 fibre network in the CNS and beyond are described. Moreover we show, in vivo, that GnRH3 is an important factor regulating GnRH3 fibre pathfinding and neurone localisation in an autocrine fashion.

Germline BRCA1 mutation and survival analysis in familial breast cancer patients in Kerala; South India.

Mutations in breast cancer susceptibility gene BRCA1 have been identified in breast or breast/ovarian cancer families from different ethnic background. We analyzed a total of 79 samples for BRCA1 mutation, using Conformation Sensitive Gel Electrophoresis (CSGE) followed by sequencing. The overall survival of BRCA1 mutation carriers was also investigated. BRCA1 mutation was detected in 11 out of the 29 (38%) patients. Four different alterations were detected of three which were novel. A missense mutation in exon 7, 465G>A was detected in 1 patient (9%). Another missense mutation 932 G>A was observed in three patients (27.3%) and a truncation mutation 1027delA, was observed in one patient (9%). The fourth type of mutation (185delAG) which also results in protein truncation was observed in 6 different patients (54.5%). Kaplan-Meier survival analysis revealed a median overall survival of 34 months for BRCA1 mutation positive breast cancer patients and 71 months for BRCA1 negative breast cancer patients. The median overall survival of BRCA1 truncation mutation carriers was 26 months. Our data showed high prevalence of BRCA1 gene mutation among breast or breast/ovarian cancer families in South India and breast cancer patients having BRCA1 mutations were associated with poor prognosis.