Kenneth D. Winkel

RelB Is Essential for the Development of Myeloid-Related CD8α− Dendritic Cells but Not of Lymphoid-Related CD8α+ Dendritic Cells

The transcription factor RelB had been shown to be important for dendritic cell (DC) development, but the type of DC involved was not clear. Here, we report that RelB mRNA is expressed strongly in CD8alpha- DEC-205- DC but only weakly in CD8alpha+ DEC-205+ DC. In addition, CD8alpha+ DEC-205+ DC are present and functional in RelB null mice, the DC deficiency being mainly in the CD8alpha- DEC-205- population. By constructing bone-marrow chimeric mice, we demonstrate that the partial deficiency in RelB null thymic DC is a secondary effect of disrupted thymic architecture. However, the deficiency in splenic CD8alpha- DEC-205- DC is a direct, stem cell intrinsic effect of the RelB mutation. Thus, RelB selectively regulates a myeloid-related DC lineage.

The Global Snake Bite Initiative: an antidote for snake bite

Clinicians have for a long time witnessed the tragedy of injury, disability, and death from snake bite that is a daily occurrence in many parts of Africa, Asia, and Latin America. To many people living in these regions, including some of the world’s poorest communities, snake bite is an ever present occupational risk and environmental hazard, an additional penalty of poverty. Like malaria, dengue, tuberculosis, and parasitic diseases, the risk of snake bite is always present. Unlike many of these other public health risks, however, the burden of human suff ering caused by snake bite remains un-recognised, invisible, and unheard by the global public health community, forgotten by development agencies and governments alike. The problem is so underrated that it was only added to WHO’s list of neglected tropical diseases in April, 2009.Yet an estimated 5·4–5·5 million people are bitten by snakes each year,

Ending the drought: New strategies for improving the flow of affordable, effective antivenoms in Asia and Africa

The development of snake antivenoms more than a century ago should have heralded effective treatment of the scourge of snakebite envenoming in impoverished, mostly rural populations around the world. That snakebite still exists today, as a widely untreated illness that maims, kills and terrifies men, women and children in vulnerable communities, is a cruel anachronism. Antivenom can be an effective, safe and affordable treatment for snakebites, but apathy, inaction and the politicisation of public health have marginalised both the problem (making snakebite arguably the most neglected of all neglected tropical diseases) and its solution. For lack of any coordinated approach, provision of antivenoms has been pushed off the public health agenda, leading to an incongruous decline in demand for these crucial antidotes, excused and fed by new priorities, an absence of epidemiological data, and a poor regulatory framework. These factors facilitated the infiltration of poor quality products that degrade user confidence and undermine legitimate producers. The result is that tens of thousands are denied an essential life-saving medicine, allowing a toll of human suffering that is a summation of many individual catastrophes. No strategy has been developed to address this problem and to overcome the intransigence and inaction responsible for the global tragedy of snakebite. Attempts to engage with the broader public health community through the World Health Organisation (WHO), GAVI, and other agencies have failed. Consequently, the toxinology community has taken on a leadership role in a new approach, the Global Snakebite Initiative, which seeks to mobilise the resources, skills and experience of scientists and clinicians for whom venoms, toxins, antivenoms, snakes and snakebites are already fields of interest. Proteomics is one such discipline, which has embraced the potential of using venoms in bio-discovery and systems biology. The fields of venomics and antivenomics have recently evolved from this discipline, offering fresh hope for the victims of snakebites by providing an exciting insight into the complexities, nature, fundamental properties and significance of venom constituents. Such a rational approach brings with it the potential to design new immunising mixtures from which to raise potent antivenoms with wider therapeutic ranges. This addresses a major practical limitation in antivenom use recognised since the beginning of the 20th century: the restriction of therapeutic effectiveness to the specific venom immunogen used in production. Antivenomic techniques enable the interactions between venoms and antivenoms to be examined in detail, and if combined with functional assays of specific activity and followed up by clinical trials of effectiveness and safety, can be powerful tools with which to evaluate the suitability of current and new antivenoms for meeting urgent regional needs. We propose two mechanisms through which the Global Snakebite Initiative might seek to end the antivenom drought in Africa and Asia: first by establishing a multidisciplinary, multicentre, international collaboration to evaluate currently available antivenoms against the venoms of medically important snakes from specific nations in Africa and Asia using a combination of proteomic, antivenomic and WHO-endorsed preclinical assessment protocols, to provide a validated evidence base for either recommending or rejecting individual products; and secondly by bringing the power of proteomics to bear on the design of new immunising mixtures to raise Pan-African and Pan-Asian polyvalent antivenoms of improved potency and quality. These products will be subject to rigorous clinical assessment. We propose radically to change the basis upon which antivenoms are produced and supplied for the developing world. Donor funding and strategic public health alliances will be sought to make it possible not only to sustain the financial viability of antivenom production partnerships, but also to ensure that patients are relieved of the costs of antivenom so that poverty is no longer a barrier to the treatment of this important, but grossly neglected public health emergency.

CD4 and CD8 expression by human and mouse thymic dendritic cells

Dendritic cells (DC) from human and mouse thymus were compared. DC from both sources were isolated by digestion with collagenase, disruption of cellular complexes with a chelating agent, selection of light density cells, immunomagnetic bead depletion of other cell types (without depletion with anti-CD4 or anti-CD8) and finally sorting for cells expressing high levels of class II MHC. Yields of DC from human and mouse thymus were comparable (around 1 DC/10(3) thymocytes), they displayed similar DC morphology, and both showed strong expression of CD11c. DC from the human thymus all expressed very high levels of CD4 but low levels of CD8. In contrast, DC from the mouse thymus expressed high levels of CD8 but only low levels of CD4. Human thymic DC were also substantially larger than mouse thymic DC. The biological significance of CD4 and CD8 expression by DC is discussed in view of this major species difference and the possibility that human thymic DC may be targets for HIV infection.

Effectiveness of snake antivenom: Species and regional venom variation and its clinical impact.

The ubiquity of venom variation in snakes poses special problems for the manufacture of antivenom and has undermined the commercial attractiveness of this class of therapeutic agent. In particular, it has been amply documented that both interspecific and intraspecific variation in venom composition can affect the neutralisation capacity of antivenoms. This may be exacerbated by the selective use of tests of venom toxicity and antivenom efficacy, such as the lethal dose and ED50, resulting in inadequate neutralisation of time, rather than dose, dependent toxins, particularly enzymes involved in defibrinogenating, haemorrhagic and necrotising venom activities. The clinical consequences can be reduced efficacy against some important venom activities or even complete treatment failure in critical envenomations. All these factors, combined with the ongoing reduction in the number of antivenom manufacturers world‐wide, and concomitant contraction in the range of available antivenoms, present significant challenges for the treatment of snakebite in the 21st century.

CARDIOVASCULAR ACTIONS OF THE VENOM FROM THE IRUKANDJI ( CARUKIA BARNESI ) JELLYFISH: EFFECTS IN HUMAN, RAT AND GUINEA-PIG TISSUES IN VITRO AND IN PIGS IN VIVO

1. We have investigated the cardiovascular pharmacology of the crude venom extract (CVE) from the potentially lethal, very small carybdeid jellyfish Carukia barnesi, in rat, guinea‐pig and human isolated tissues and anaesthetized piglets.

Immunological and Toxinological Responses to Jellyfish Stings

Just over a century ago, animal responses to injections of jellyfish extracts unveiled the phenomenon of anaphylaxis. Yet, until very recently, understanding of jellyfish sting toxicity has remained limited. Upon contact, jellyfish stinging cells discharge complex venoms, through thousands of barbed tubules, into the skin resulting in painful and, potentially, lethal envenomations. This review examines the immunological and toxinological responses to stings by prominent species of jellyfish including Physalia sp (Portuguese Man-o-War, Blue-bottle), Cubozoan jellyfish including Chironex fleckeri, several Carybdeids including Carybdea arborifera and Alatina moseri, Linuche unguiculta (Thimble jellyfish), a jellyfish responsible for Irukandji syndrome (Carukia barnesi) and Pelagia noctiluca. Jellyfish venoms are composed of potent proteinaceous porins (cellular membrane pore-forming toxins), neurotoxic peptides, bioactive lipids and other small molecules whilst the tubules contain ancient collagens and chitins. We postulate that immunologically, both tubular structural and functional biopolymers as well as venom components can initiate innate, adaptive, as well as immediate and delayed hypersensitivity reactions that may be amenable to topical anti-inflammatory-immunomodifier therapy. The current challenge for immunotoxinologists is to deconstruct the actions of venom components to target therapeutic modalities for sting treatment.

Comparative proteomic analysis of the venom of the taipan snake, Oxyuranus scutellatus, from Papua New Guinea and Australia: Role of neurotoxic and procoagulant effects in venom toxicity

The venom proteomes of populations of the highly venomous taipan snake, Oxyuranus scutellatus, from Australia and Papua New Guinea (PNG), were characterized by reverse-phase HPLC fractionation, followed by analysis of chromatographic fractions by SDS-PAGE, N-terminal sequencing, MALDI-TOF mass fingerprinting, and collision-induced dissociation tandem mass spectrometry of tryptic peptides. Proteins belonging to the following seven protein families were identified in the two venoms: phospholipase A(2) (PLA(2)), Kunitz-type inhibitor, metalloproteinase (SVMP), three-finger toxin (3FTx), serine proteinase, cysteine-rich secretory proteins (CRISP), and coagulation factor V-like protein. In addition, C-type lectin/lectin-like protein and venom natriuretic peptide were identified in the venom of specimens from PNG. PLA(2)s comprised more than 65% of the venoms of these two populations. Antivenoms generated against the venoms of these populations showed a pattern of cross-neutralization, corroborating the immunological kinship of these venoms. Toxicity experiments performed in mice suggest that, at low venom doses, neurotoxicity leading to respiratory paralysis represents the predominant mechanism of prey immobilization and death. However, at high doses, such as those injected in natural bites, intravascular thrombosis due to the action of the prothrombin activator may constitute a potent and very rapid mechanism for killing prey.

Ant sting mortality in Australia.

We investigated ant sting related fatalities in Australia over the period 1980-1999. Data was obtained from the Australian Bureau of Statistics and state coronial authorities. Six ant sting-related fatalities were identified, five in Tasmania and one in New South Wales. All were males aged between 40 and 80-years-of-age and most (5/6) had prior histories of jumper or bull ant (Myrmecia spp.) venom allergy. However, none of the deceased carried injectable adrenaline and most died within 20 min of a single sting. Significant cardiopulmonary co-morbidities were identified in all cases and, in addition, moderate-severe laryngeal oedema and coronary atherosclerosis was observed in most (4/6) cases at autopsy. Where ascertained, Myrmecia ant venom specific immunoglobulin E antibodies levels were always elevated and fell into two distinct patterns of immunoreactivity. Adult Tasmanian males with a prior history of ant venom allergy and cardiopulmonary co-morbidities are therefore at highest risk of a fatal outcome from ant stings. Deaths may be avoided by the early recognition of anaphylaxis and self-treatment with adrenaline as well as by the development of purified Myrmecia ant venom immunotherapy.

Marine Stingers: Review of an Under-Recognized Global Coastal Management Issue

Dangerous marine stingers (jellyfish) are an emotive issue in tropical Australia, where they are widely regarded as the number one marine health threat. However, numerous severe and fatal stings have been reported throughout the tropical and temperate seas of the world, indicating that marine stingers are a global health problem. Further, life-threatening jellyfish stings are more frequently reported globally now compared to earlier decades, possibly as a result of improved recognition and reporting, or increased spatial and/or temporal distribution or densities. As stinging incidents may also have significant financial implications (lost tourism revenues and liability settlements), and the treatment of envenomed patients comes at high cost to the taxpayer, this issue is also a management challenge. This article outlines suggested approaches, based largely on Australian experience, for dealing with this under-recognized global coastal management issue.