The ongoing evolution of humanness: perspectives from Darwin to de Chardin
105, November/December 2009

J S Buckeridge email@example.com
School of Civil
Environmental & Chemical Engineering
RMIT University
GPOBox 24763001VICAustralia

The ongoing evolution of humanness: perspectives from Darwin to de Chardin
105, November/December 2009C693BB87F0A33628363A5301A03B64A6
However an ability to reason or to possess nous is not restricted to H. sapiens-and it is often in adversity that the most interesting challenges to this presumption are made.During the tragic forest fires that ravaged Victoria, Australia in February 2009, 173 humans and an estimated million 'large' animals died (Christine  Nixon, pers.comm.).Some remarkable accounts of survival and interspecies interaction were made during the 2009 Victorian Bushfires Royal Commission hearings held in Melbourne in the months following the fires.In one, two survivors related that, in the face of an advancing wall of flames, they sought safety in a short, open culvert.Shortly after entering this, they were joined by a large eastern grey kangaroo, which remained in very close proximity to them until it concluded the danger had passed.At that point it hopped out and made its way through the cooling embers to safety.The fire front was recorded to pass through areas in about 30 minutes, so we may conclude that two humans and one large wild animal shared a small space for about that time.It is most unlikely that any wild kangaroo would approach humans under normal conditions.What happened here is remarkable, as the kangaroo clearly observed danger, was able to seek refuge, and although it would certainly have been aware that it was not alone in the drain, elected to stay there with animals that would normally be a threat to it until it determined outside conditions had returned to a safe level.A bold deduction would be that this was an excellent demonstration of a kangaroo exercising utilitarianism, but I could not possibly state this in a scientific journal.A further example of interspecies interaction during the fires involved a man leaping into a dam.He swam to the central, deepest part, where he repeatedly submerged

Introduction

Palaeontology and palaeobiology are sciences that deal with the history of past life.In particular they use the disposition, the remains and the impressions of fossils to reconstruct the phylogeny of taxa, their biogeographic patterns and the types of environments in which they once lived.On the whole, they deal with concrete concepts.Thus, an evaluation of humanness, a concept with no apparent fossil record, is a pursuit that one would not expect of palaeontologists.Nonetheless, the journey to understand and define character(s) of that phenomenon we call 'man' has felt the tread of some of our greatest palaeontologists, including Charles Darwin and Teilhard de Chardin.The objective of this paper is to explore several of the paths in mankind's quest to understand the nature of 'being human'-the thesis being that if we do not know what we are, our ability to plan for the future is severely compromised.These two scientists are specifically chosen in this essay in recognition of the manner in which their theses so strongly challenged contemporary science, religion and philosophy.

Pursuing 'humanness'

In a simplistic way, humanness may be defined as being aware; indeed, Linnaeus embodied this concept as recently as 1758, when he coined Homo sapiens, or 'thinking man' to systematically define us.Much earlier, the philosopher Aristotle (384-322 BC) sought to define the very nature of the humanness that he saw as the end-purpose for humanity.The deductive path that Aristotle follows (below) illustrates that, although we share many features with other higher animals, it is an ability to 'reason' that distinguishes us:

The human function: What, then, could this be?For living is apparently shared with plants, but what we are looking for is the special function of a human being; hence we should set aside the life of nutrition and growth.The life next in order is some sort of life of sense-perception; but this too is apparently shared, with horse, ox and every animal.The remaining possibility, then, is some sort of life of action of the [part of the soul] that has reason… The human function is the soul's activity that expresses reason [as itself having reason] or requires reason [as obeying reason].

The nature of humanness is discussed from observations made by himself to keep cool and to douse the falling cinders.During the period of greatest conflagration he was tightly surrounded by a number of ducks, most which were within touching distance.

It would be arrogant indeed to conclude that the actions of the kangaroo and birds were simply of instinct, and therefore fail to demonstrate any higher level of nous.

Although ability to rationalise is clearly not the prerogative of humans, it is nonetheless a capacity that is not held equally in other species.Rather, it is a progressive faculty, particularly within mammals, such that the ability to reason in different species may be arranged in a continuum. 2ther traits that must be assessed in a journey to define humanness include the development and sustainability of complex social behaviour; the ability to produce and appreciate art and music; the appreciation of beauty; spatial and temporal awareness; the development of complex oral and written language; religion; and not least, the moral virtues, 44 of which are: assertiveness, beneficence, benevolence, cleanliness, commitment, compassion, confidence, cooperation, courage, courtesy, creativity, detachment, diligence, enthusiasm, excellence, flexibility, forgiveness, generosity, gentleness, honesty, honour, humility, idealism, joyfulness, justice, love, loyalty, moderation, modesty, orderliness, patience, peacefulness, perseverance, purposefulness, respect, responsibility, self-discipline, service, tact, thankfulness, tolerance, trustworthiness, understanding, unity.

The virtues, espoused by Aristotle, are derived the Greek DgJZ (arete); a term encompassing goodness and excellence, wherein any choice made is one that lies between a deficiency and an excess.For example, courage is the mean between cowardice and foolhardiness.Any one of the above virtues alone does not define good, and it follows that practice of any in isolation cannot be seen as manifestly human-and may not even be moral.Whether animals are capable of moral judgement is unknown, although there is some evidence of this behaviour in primates, social carnivores and some cetaceans. 3Ironically, much of the journey to human dominance over the biosphere has been achieved through exercising traits that are clearly not in the least virtuous, although they may, in the short term, illustrate survival of the fittest, e.g.genocide and waging of war.The manner in which we address mankind's greatest crime, environmental degradation, will ultimately decide which species is the fittest.On past record, it may not necessarily be H. sapiens.

Returning to the earlier traits, humanness defined by the possession of social behaviour that permits coexistence in densely populated communities is flawed, as this character is demonstrated by many species, including invertebrates such as bees and ants.An appreciation of music, especially the creation of music, is sometimes cited as a uniquely human trait; however, there are numerous examples of song created by birds and cetaceans.Birds in particular are capable of mimicking human music, just as we do with birdsong.Music definitely has a calming effect-on humans, and also on many other higher animals.Farmers in many countries use classical music to placate cows during milking.However, when it comes to animals making music, there is still debate about the purpose-whether it is functional, or for pleasure, and whether we are simply trying too hard to anthropomorphise. 4 A distinction humans do have however, is the ability to inscribe the music, e.g. in the written form, or digitally.

As to beauty, the concept, and appreciation, of it entertained Aristotle, 5 and Darwin deliberated long on its significance in human evolution-and concluded that an awareness of beauty within the environment, as opposed to a response to beauty as a sexual attractant (as in many animals), was a trait refined and acquired in humans through culture.We should remember however, that Darwin was a keen observer of nature, and in The Descent of Man, and Selection in Relation to Sex (hereafter Descent of Man) he notes that some animals deliberately collect bright objects with which to adorn their nests. 2Thus an appreciation of beauty in itself is not useful in defining humanness.

Nor is an appreciation of human language restricted to humans, for example, that most domesticated of species, Canis familiaris is certainly capable of understanding not just nuances, but simple words.Many dogs can comprehend the spelled word, e.g.'w-a-l-k' and respond accordingly (Fig. 1).A sceptic may state that dogs do not understand words-rather they are just very good at reading human body language.Indeed this may be so, but also so for human infants.In language then, it is perhaps only our ability to manipulate symbols in a complex manner that is distinctly human.Some animals have demonstrated an ability to count, 6 and most animals have an awareness of space, although an appreciation of the relationship between time and geography is much more difficult to assess.Many animals migrate: they clearly know where to go, and when, although this is for the most instinctive rather than cognitive.Although we currently have no way of knowing exactly what animals are capable of thinking of, it is perhaps the human ability to think beyond the realms of reality, to model present trends and hypothesise on both the distant past and the future that is distinctive.Nonetheless, this does not significantly reduce the elusiveness of the concept of humanness.

Until the inception of the Enlightenment in Western Europe at the beginning of the 18th century, man was generally considered distinct from the biosphere.This Judaeo-Christian view has humans created separately from all other living organisms, being differentiated by creation in the image of God.Not surprisingly, man was given dominion over nature.This was to change in the mid-1800s, when one philosopher in particular, Charles Darwin, took a different turn when he ventured to suggest that humans were part of a continuum with other animals, rather than the product of any special creation. 2,7olution and the emergence of human culture Whilst the preceding section pursued, and in part assessed the essence that defines humanity, the following reviews the climate of aversion that had, up to the mid-19th century, obstructed the placement of H. sapiens-with respect to all other living organisms -into the scheme of origins through natural selection.

Review Articles

Man's journey away from immortality

As a young graduate student I was very much preoccupied with the phylogeny, evolution, and geographic and temporal distribution of barnacles.This provided an extraordinary opportunity to research the group of organisms that had, some 130 years earlier, sufficiently inspired Charles Darwin for him to devote more than eight years to their study.However, it is how he interpreted the barnacles that is relevant here-for in four volumes ranging over 1851-1854, the barnacles were described in an order that we would find acceptable in today's systematic taxonomy. 8,9Darwin began his study of barnacles (or cirripedes) after finding a minute, rather unusual, burrowing cirripede in the shell of the South American gastropod Concholepas.His dissection and description of this taxon aroused his natural curiosity of the whole group, which had only recently been recognised as crustacean rather than molluscan.Cirripedes were, at that time, not well understood, but a growing amount of excellent fossil material, supplemented by collecting considerable numbers of recent specimens, provided an irresistible opportunity to unravel the taxonomic relationships of the group. 9Finally, it was the encouragement of John Edward Gray, then Keeper of Zoology at the British Museum, that ensured his long dedication to the group. 10It is reasonable to conclude that it was the cirripedes that provided Darwin with the insight into how natural selection functioned.Surprisingly, he referenced them little in his seminal work On the Origin of Species by Means of Natural Selection (hereafter Origin of Species), although it was his barnacle research that confirmed his status as a biologist, and for this he was awarded the Royal Society's Royal Medal in 1853. 9arwin was somewhat reluctant to publish his views on humanity's place in evolution, and this probably reflects his concern with public acceptance, or otherwise, of his 'species theory' .Although 'enlightened' with respect to the previous century, Britain in the mid-to late-19th century was not the secular society it is today; further, it is important to reflect upon the very considerable influence that the established church (i.e. the Church of England) had at that time-in all aspects of life, including science.Darwin was all too aware of this, and agonised over the polarising effect he knew his work would have-as many saw it as a direct threat to the widely accepted and socially cohesive relationship between man and God.In light of strong criticism, he did remove one significant impediment to wider acceptance of his work by including suitable reference (and reverence) to a Creator in the closing sentences of later editions of the Origin of Species. 11,12lthough he had planned to include reference to human beings in his 1868 book, The Variation of Animals and Plants under Domestication, he did not do so, apparently because the size of this work, which, as two volumes, ended up much greater than he had originally planned. 13It was ultimately in 1871, more than ten years after the publication of Origin of Species, that Darwin was sufficiently emboldened, or believed his Origin of Species was sufficiently well accepted, for him to publish The Descent of Man (Fig. 2).

The key tenet in the Descent of Man is that H. sapiens is descended from some less highly organised (= evolved) form.Amongst the evidence for this that Darwin cites: • A shared embryonic development.

• A shared morphology (e.g. the structure of the skull, limbs and musculature, especially with other mammals).

• Geological succession (i.e. the fossil record).Although he acknowledges that there are individual differences between man and other animals, Darwin concluded that any perceived differences would have been induced by the same general causes and therefore must obey the same laws (i.e.those of natural selection) as the lower animals.He takes great effort to emphasise this principle:

The great principle of evolution stands up clear and firm… it is incredible that all these facts should speak falsely.He who is not content to look, like a savage, at the phenomena of nature as disconnected, cannot any longer believe that man is the work of a separate act of creation.

One area where Darwin does see distinction between humans and other animals involves the level of intellect and the moral disposition of humans.He acknowledges that these mental powers exist in higher animals, and that they are of the same nature; but he concludes that the difference is simply the degree to which they are developed.

However, Darwin views the principal distinction between man and animals as religious: with the special essence that defines humans being the impact of a deity.He was confident that humans were spiritual creatures, and that belief in 'spiritual agencies' was ubiquitous in human societies; however, it may well follow from this that an instinctive belief in God is the self-fulfilling argument for God's existence.

The belief in God has often been advanced as not only the greatest, but the most complete of all the distinctions between man and the lower animals.It is however impossible, as we have seen, to maintain that this belief is innate or instinctive in man.

That Darwin was unable to resolve whether or not a beneficent God arose in humanity as a direct result of the advance of culture is demonstrated by his apparent agnosticism at his death.More than a century later, this issue remains unresolved in science.

South African links

In 1836, during the voyage of HMS Beagle, Darwin visited South Africa and stayed several weeks at Simon's Bay, Cape Town. 14Although we are not able to differentiate FitzRoy's words from those of Darwin's, the comments they made at that time in a letter to the South African Christian Recorder are relevant here because they give us an insight into Darwin's worldview. 15In the process of writing in defence of land acquisition by missionaries, they extolled the elevated place of Europeans, especially English gentlemen and Church of England missionaries, who, they asserted, began the civilising process of native peoples through the teaching of the gospel.Although Darwin lived at a time when there were widespread and very different perceptions of the tribes of the human race to those we currently have, it is clear from his abhorrence of slavery that he was deeply disturbed by the inequity and harsh treatment that much of humankind received.

His views on women too are, from a 21st-century perspective, politically unacceptable-even though he was viewed as a seditious liberal by many of his countrymen.Again this must be seen in context of his times, for the resolution of universal suffrage for women was still many decades away.Charles Darwin's perceptions were a reflection of 19th-century morality.Nonetheless, I cannot even begin to appreciate how his capable and indefatigable wife Emma would have viewed his observation that:

Man is more courageous, pugnacious, and energetic than woman, and has a more inventive genius.

Reconciling the religious-science nexus

The French Jesuit palaeontologist, Pierre Teilhard de Chardin was born in 1881, one year before Darwin's death; like Darwin, de Chardin was to visit South Africa.He did so twice-in 1951 and 1953, inspecting the Pliocene Australopithecus africanus caves at Sterkfontein in the company of the distinguished South African hominid palaeontologist John T. Robinson. 16De Chardin's remarkable career was by this time almost over; although he made a significant contribution to anthropology, this was not achieved in Paris or Rome, for he had spent most of his adult life living in exile overseas.His formative years as a priest were in Europe, but his notions of life, although germinated in youth, were to come to fruition in China where he worked for two decades and where, amongst other extraordinary achievements, he was co-discoverer of Peking man.(He was also 'co-discoverer ' of the now infamous Piltdown man, found in an English quarry, which was subsequently demonstrated to be a hoax 17 ).De Chardin's most widely known work, The Phenomenon of Man, completed in 1938, was not published until 1955, shortly after his death. 18In this book, de Chardin the scientist confirms his acceptance of the antiquity of the Earth and the progression of life, from the simplest forms to the most complex, as reflected in the fossil record.He is a Darwinist.However, this science is somewhat compromised by mysticism when introducing the ubiquitousness of spirituality-a concept with which, he is confident, we must all be most familiar. 18Although de Chardin is forthright in recognising that spirituality is opaque scientifically, human spiritual evolution develops as the more profound aspect of de Chardin's polemic.It is apparent from the outset that The Phenomenon of Man was intended as a scientific, rather than a theological work.Nonetheless, it is a work that many scientists would not recognise as science-this book is not founded in empiricism, although this should not detract us from the message.De Chardin seeks to unravel the greatest question humanity faces: Who are we, and what is our purpose?As noted, Aristotle saw the end point of being human as the exercise of rational capacities like excellence and intelligence; he concluded that these are generally best developed within the context of friends, or community. 1To de Chardin, however, the end point for humanity (hominisation) was still some considerable way off.

De Chardin believed that all species mutate: they either evolve through time or become extinct.Of course this process also includes humans.He saw the end point of human evolution as the formation of the noosphere, a term coined to describe an invisible layer of thought that would surround the Earth, and which would equate to the sum total of humanity's mental and spiritual state. 18In his noosphere, all human culture, love and knowledge would be subsumed into a shared mental and spiritual state.This would follow the evolution of a more self-reflective humanity, and would end at what he called an ' omega point' .He deemed every individual as possessing a vital role in the evolution of the world, with each, through cooperative effort, being clearly cognisant of his or her individual task in achieving the ' omega point' .

De Chardin's premonition of a global web through establishment of the noosphere has, in a way, been realised much earlier than he would have anticipated-in the worldwide web.However, the breadth and magnitude of this web is probably not what de Chardin would have anticipated, or have wished.As he would have hoped, the worldwide web does constitute a medium in which individuals may express their potential (online) to the full, and it also represents, to an increasing degree, an accumulation of much of human expression.However, it cannot currently be categorised as a vehicle that is advancing humanity to a spiritual communion with a personal God.Importantly, it is through the latter that De Chardin saw fulfilment of his noosphere.

De Chardin's work attempts to weave together both science and a deep Christian mysticism in a manner imbued with optimism and anticipation.His argument is often difficult to follow, especially if viewed from a scientific perspective; as a rational, empirical scientific thesis then, it fails-but as a reflection on the potential for humanity to reach nirvana, it is a truly wondrous success.

Drawing the threads together

The way in which humans perceive themselves is, in the main, the substance of literature-the written word thus provides a cogent reflection of the nature of a particular society at a fixed point in time and place.This reflection comes to us not simply through the art of poetry and prose, but through the utilitarian languages of commerce, engineering, law and science.However, there is another less rigid usage, wherein language defines those quintessential human genres of satire, irony, mockery, censure and humour.Indeed, it is through the use of these that we may gauge the level (and depth) of a civilisation.For example, as the first decade of the new millennium draws to a close, humanity is at last becoming painfully aware of the outcomes of both its success and its excess.Faced with what is an apparently inexorable and imminent global warming, and an intransigent Intergovernmental Panel on Climate Change that is adamant that the change is driven primarily by anthropogenic carbon dioxide, it is more than refreshing to read a novel and moderately well-argued satirical dissertation that claims that a strong causal pathway exists between global warming and Aztec blood sacrifice rituals, 19 rather than carbon dioxide levels.The use of satire and humour to probe, to expose and to question all in the pursuit of truth is uniquely human.

On a more sobering note, the 16th-century writings of the poet William Shakespeare, challenge us to conclude that, of the many human traits we possess, one has primacy in separating humans from other animals, and that is the concept of mercy.Shakespeare argues that it is through exercise of mercy that one adopts a godly trait, and thus demonstrates the best in a human.Importantly, it is this approximation of behaviour to that of a (beneficent) deity that is used to demonstrate the highest good, i.e. the quality of human action is judged on the basis of how closely the activity approximates to those anticipated of God.

The quality of mercy is not strain'd, It droppeth as the gentle rain from heaven Upon the ground beneath: it is twice bless'd; It blesseth him that gives and him that takes: 'Tis mightiest in the mightiest: it becomes The throned monarch better than his crown; His sceptre shows the force of temporal power, The attribute to awe and majesty, Wherein doth sit the dread and fear of kings, It is an attribute to God himself; An earthly power doth then show likest God's When mercy seasons justice.

Last words

Wherein these last words, the essence of 'being a good human' is determined and a rationale for religion, and the order that this may inspire, is entertained.

Humans are social animals.This in itself is not the essence of humanness, as many other species are social, or colonial.What is important about the social behaviour of humans is the manner in which society has evolved, and the mechanisms through which social cohesion are achieved.Edward Wilson, who coined the term sociobiology, recognised the uniqueness of human language, with its intricate symbolism, quality and grammar. 21ilson also reminded us that humans are remarkably easy to indoctrinate.Further, he concluded that although this intellectual plasticity has many advantages, if uncontrolled it would lead to social anarchy.He thus saw the evolution of religion as the antidote to this plasticity. 21How Darwin would have viewed Wilson's conclusions is somewhat intangible; in his early years Darwin was a theist, and even at the end he was agnostic rather than an atheist. 22Darwin did not close the door on religion, although he clearly entertained the idea that God (the Creator) was potentially a manifestation of human culture:

The idea of a universal and beneficent Creator of the universe does not seem to arise in the mind of man, until he has been elevated by long-continued culture.

Darwin's understanding of the evolution of spiritual awareness in conjunction with the evolution of community, continues the link evidenced in the works of Aristotle (and de Chardin), namely that humans are best able to reach true happiness through positive interaction with friends and community. 1 an interesting parallel, recognition of the significance of community lies within the Bantu concept of ubuntu, a process that seeks to establish unity through the act of living together as harmonious communities. 23The essence of ubuntu is the realisation that a man is not an island, and that a single human individual cannot reach full potential in isolation.

Rather, for better or for worse, we are the products of our community.A form of the global community, and global communication, that de Chardin envisaged is now with us.How we exercise the knowledge this system provides will determine the future of our species.

The author wishes to thank the two anonymous referees who made useful comments on an earlier draft of the manuscript.

Fig. 1 .
1
Fig. 1.To think is to know.However, knowing is not an exclusively human characteristic.(Canis familiaris, alias Buster the boxer.)

Fig. 2 .
2
Fig. 2. The title page from the first edition of Darwin's The Descent of Man and Selection in Relation to Sex (1871, John Murray, London).

Aristotle in 4th-century Greece, through to those of Charles Darwin, Teilhard de Chardin and William Shakespeare. Attempts to define humanness upon a narrow range of criteria, as some have tried, is argued as flawed, for humanness is more elusive than a single or a few demonstrated phenomena. The path that Darwin pursued in determining the place of humans in nature in his book The Descent of Man, and Selection in Relation to Sex is assessed from a 19th- century perspective; the difficulties he faced, both personally and with the broader public, are reviewed and then evaluated in a modern context. Darwin's thesis adheres to scientific principles, and is debated, defended and later verified on these principles. This is somewhat at variance to the approach adopted by the priest-scientist de Chardin a century later in his major work, The Phenomenon of Man-in which an attempt is made to reconcile a deep Christian faith with science. De Chardin scores well from a theological viewpoint, but fails on scientific grounds as his thesis moves beyond the realms of empiricism into mysticism. Surprisingly, de Chardin's predicament of a future wherein human evolution enters a new stage of consciousness through the noosphere (an invisible layer of thought encompassing the globe) has been partially realised through the worldwide web, although the nature of the web is almost certainly not what de Chardin might have anticipated, or desired. Science too fails to answer all, particularly the nature of God. Darwin considered the Creator in several of his works and does not dismiss the concept of a farseeing deity, although we are left with the notion that he died agnostic. Humanness is derived from an elevated moral code and this is reflected in our arts, particularly literature, wherein we may temporally reflect upon quintessential human traits such as mercy. However, expression of the arts is only achievable by the individual being part of a greater whole: the human community, the essence of which is distilled in the Bantu concept of ubuntu, wherein humanness is best realised through the act of living in harmony. Key words: human evolution, Charles Darwin, Teilhard de Chardin, culture, morality, virtues

Received 27 August. Accepted 11 November 2009.
South African Journal of Science 105, November/December 2009

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Potential 'universal' blood test for cancer discovered
28 July 2014

Potential 'universal' blood test for cancer discovered
28 July 2014
Researchers from the University of Bradford, UK, have devised a simple blood test that can be used to diagnose whether people have cancer or not.

The test will enable doctors to rule out cancer in patients presenting with certain symptoms, saving time and preventing costly and unnecessary invasive procedures such as colonoscopies and biopsies being carried out. Alternatively, it could be a useful aid for investigating patients who are suspected of having a cancer that is currently hard to diagnose.

Early results have shown the method gives a high degree of accuracy diagnosing cancer and precancerous conditions from the blood of patients with melanoma, colon cancer and lung cancer. The research is published online in the FASEB Journal, the US Journal of the Federation of American Societies for Experimental Biology.

The Lymphocyte Genome Sensitivity (LGS) test looks at white blood cells and measures the damage caused to their DNA when subjected to different intensities of ultraviolet light (UVA), which is known to damage DNA. The results of the empirical study show a clear distinction between the damage to the white blood cells from patients with cancer, with pre-cancerous conditions and from healthy patients.

Professor Diana Anderson, from the University's School of Life Sciences led the research. She said: "White blood cells are part of the body's natural defence system. We know that they are under stress when they are fighting cancer or other diseases, so I wondered whether anything measureable could be seen if we put them under further stress with UVA light. We found that people with cancer have DNA which is more easily damaged by ultraviolet light than other people, so the test shows the sensitivity to damage of all the DNA -the genome -in a cell."

The study looked at blood samples taken from 208 individuals. Ninety-four healthy individuals were recruited from staff and students at the University of Bradford and 114 blood samples were collected from patients referred to specialist clinics within Bradford Royal Infirmary prior to diagnosis and treatment. The samples were coded, anonymised, randomised and then exposed to UVA light through five different depths of agar.

The UVA damage was observed in the form of pieces of DNA being pulled in an electric field towards the positive end of the field, causing a comet-like tail. In the LGS test, the longer the tail the more DNA damage, and the measurements correlated to those patients who were ultimately diagnosed with cancer (58), those with precancerous conditions (56) and those who were healthy (94).

"These are early results completed on three different types of cancer and we accept that more research needs to be done; but these results so far are remarkable," said Professor Anderson. "Whilst the numbers of people we tested are, in epidemiological terms, quite small, in molecular epidemiological terms, the results are powerful. We've identified significant differences between the healthy volunteers, suspected cancer patients and confirmed cancer patients of mixed ages at a statistically significant level of P
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Direct-writing of PbS nanoparticles inside transparent porous silica monoliths using pulsed femtosecond laser irradiation

Abdallah Chahadih
Hicham El Hamzaoui
Rémy Bernard
Luc Boussekey
Laurence Bois
Odile Cristini
Marc Le Parquier
Bruno Capoen
Mohamed Bouazaoui
Direct-writing of PbS nanoparticles inside transparent porous silica monoliths using pulsed femtosecond laser irradiation
N A N O E X P R E S S
Pulsed femtosecond laser irradiation at low repetition rate, without any annealing, has been used to localize the growth of PbS nanoparticles, for the first time, inside a transparent porous silica matrix prepared by a sol-gel route. Before the irradiation, the porous silica host has been soaked within a solution containing PbS precursors. The effect of the incident laser power on the particle size was studied. X-ray diffraction was used to identify the PbS crystallites inside the irradiated areas and to estimate the average particle size. The localized laser irradiation led to PbS crystallite size ranging between 4 and 8 nm, depending on the incident femtosecond laser power. The optical properties of the obtained PbS-silica nanocomposites have been investigated using absorption and photoluminescence spectroscopies. Finally, the stability of PbS nanoparticles embedded inside the host matrices has been followed as a function of time, and it has been shown that this stability depends on the nanoparticle mean size.

Introduction

New directions of modern research have emerged during the last decade, which has broadly been defined as nanoscale science and technology [1,2]. These new trends involve the ability to fabricate, characterize, and manipulate artificial structures, the features of which are controlled at the nanometer scale. The semiconductor properties of lead sulfide PbS have widely been used in elements such as detectors operating in the infra-red spectral region (from 850 to 3100 nm) [3], solar batteries [4], and advanced optoelectronic devices [5]. In most of recent applications, useful properties of the PbS nanoparticles arise from the strong quantum confinement effect of the charge carriers (Bohr radius of 18 nm) and the associated optical nonlinear property [6].

Semiconductors may be included in a variety of media including polymers [7][8][9], micelles [10,11], glassy particles [12], and zeolites [13]. The synthesis of semiconducting nanoparticles in glass matrices has become very important. A number of methods to synthesize semiconductor nano-crystals have been employed and these include hydrothermal synthesis [14], chemical bath [15], spray pyrolysis [16], laserheated evaporation [17], combustion synthesis [18], and the sol-gel technique [19]. The sol-gel technique has become very popular because of the high chemical homogeneity of the products, also to the low processing temperatures, the possibility of controlling the particles size and morphology. The sol-gel-derived materials provide excellent matrices for a variety of organic and inorganic compounds. Silica porous matrices can be obtained using the sol-gel process and depending on how the wet gel is dried, the porosity of the materials can be tailored [20]. Such a controlled porosity can be used to dope it with active elements by impregnation with appropriate solutions.

A few articles have been reported in the literature on the crystallization of PbS nanoparticles by heat-treatment inside a silica matrix prepared by sol-gel method [21][22][23]. In this case, however, the precipitation of semiconductor nanoparticles has been realized without any space-localization. To reach this aim, the use of a laser irradiation is a promising method. Recently, local precipitation of PbS nanoparticles in melting glass matrix has been reported [24,25]. Chao et al. [24] used pulsed femtosecond laser irradiation (800 nm, 1 kHz, and 184 fs) followed by a heat treatment at a temperature of 450°C to precipitate PbS nanoparticles inside irradiated areas of a melted glass. Takeshima et al. [25] reported the local precipitation of PbS inside a melted glass without annealing using a pulsed femtosecond laser irradiation at high repetition rate (250 kHz, 200 fs, 750 mW). Recently, we have reported the space-selective growth of CdS nanoparticles, at ambient conditions, inside silica xerogels by laser irradiation [26].

Herein, we report on our investigations, based on a pulsed femtosecond laser irradiation performed with a low repetition rate in ambient conditions, for the localized growth of PbS nanoparticles inside a deep volume of porous silica monoliths. Moreover, we provide a study on the stability of PbS nanoparticles as a function of time.

Experimental

Porous silica xerogels with a thickness of 2 mm has been prepared by a sol-gel route using tetramethylorthosilicate [20]. The obtained silica monolith exhibited interconnected pores of mean diameter 5.4 nm, a specific surface area of 360 m 2 g -1 and a pore volume of 0.49 cm 3 g -1 . All these parameters were determined by isothermal nitrogen sorption measurements, as previously described [20]. The obtained porous silica samples were impregnated for 4 h into an aqueous solution (S) composed of the lead acetate as a lead precursor and of thiourea as a sulfur precursor. The concentration of each precursor in S was 0.37 mol L -1 and the molar ratio between Pb and S sources was kept to 1.

The irradiation experiments were performed using a Ti:sapphire oscillator, followed by a regenerative amplifier producing 120 fs pulses at 800 nm with a 1 kHz repetition rate. The laser beam, with an average power ranged between 10 and 40 mW, was focused through a 10× microscope objective with a numerical aperture of 0.30. The obtained spot, of a diameter estimated to 2 μm, was located inside the volume of the silica matrix and scanned laterally at a rate of 1 mm s -1 . A tight network of irradiated lines with a step size of 20 μm has been performed to cover a wide zone. The colored area was then large enough to be observed with the naked eye and to be optically characterized. Figure 1 of the Sloaded silica monolith shows the irradiated area inside the deep volume of the xerogel for a laser power of 10 mW, corresponding to a crest irradiance of about 2.5 × 10 15 W cm -2 .

Absorption spectra were recorded at room temperature in the irradiated zone using a Perkin-Elmer lambda 19 UV-Vis-IR double beam spectrometer.

Micro-photoluminescence measurements have been performed using a HORIBA Jobin Yvon IHR320 spectrometer coupled with a microscope at room temperature. Both 351.1 and 514.5 nm laser lines of an argon laser have been used to excite the PbS-doped silica matrices in the fs-irradiated zone. The incident laser power was kept low enough to avoid any further precipitation of PbS nanoparticles.

XRD patterns are recorded on a Philips X'Pert diffractometer equipped with a monochromator, using Cu Kα radiation.

TEM characterization was performed on a microscope FEI Tecnai G2-20 twin with a 200 kV acceleration voltage. For TEM imaging, the precipitated zone of a doped sample was grinded into a powder and then deposited onto a copper grid previously coated with a thin carbon membrane. The powder was then metalized with a vaporized carbon layer.

Result and discussion

The irradiation wavelength 800 nm corresponds to energy much smaller than the band gap of silica. Hence, no linear absorption can occur when the matrix is irradiated, thus allowing the laser beam to penetrate deeply inside the material. Hence, it is possible to embed a PbS layer of nanoparticles anywhere in the monolith. However, even at the lowest average power corresponding to tremendous laser peak power, a multi-photon absorption exists. As a result, the impact area is highly localized, confined within the focal volume of the focused beam. The irradiated zone showed brown or black color, depending on the average power of the laser beam. This color is easily visible to the naked eye, as shown in Figure 1.

The formation of PbS nanocrystals inside the porous silica glass, after laser irradiation, was confirmed by different analysis techniques. According to the different analysis techniques, we have irradiated at different depths: just under the surface for XRD measurements and for the other characterisations, the focused beam was in the middle of the monolith. TEM analysis has been performed to evidence the formation of PbS nanoparticles. Figure 2a shows a TEM image of the irradiated area at 40 mW. Quasi-spherical nanoparticles, with a size distribution ranging between 5 and 12 nm, have been observed. Figure 2b shows the EDX analysis recorded on the same area as in Figure 2a Optical absorption spectra have been recorded in the irradiated and in the non-irradiated areas, as shown in  Figure 3(left). The absorption spectrum of the zone irradiated with an average power of 40 mW is red-shifted as compared to the absorption spectra of the non-irradiated zone. Moreover, one can note that the absorbance baseline in the irradiated area is much higher than those of the non-irradiated zone. Such a strong absorption level has been attributed to the formation of PbS nanoparticles in high concentration. The absence of any excitonic absorption structure in the absorption spectra may have two main reasons: first, the weak exciton binding energy because of the strong Coulomb screening in narrow gap semiconductors and second, the existing size distribution of the nanocrystallites.

Photoluminescence spectra of the area irradiated with a power of 40 mW and in the unirradiated area are shown in Figure 3(right). Under excitation with the 351 nm laser line, the non-irradiated area emits light in a large band centered on 600 nm, while the excitation of the irradiated area at the same wavelength results in two emission bands: the first one being centered on 650 nm and the second one around 910 nm. Consequently, the bands peaking at 600 and 650 nm could be attributed to the luminescence of PbS precursors and SiO 2 host matrix, since the S-doped silica absorption at 350 nm is still strong. On the other hand, the PL band centered at 910 nm is due to the PbS nanoparticles. To confirm our assumption, the same PL spectra were recorded under excitation at 514.5 nm. At this wavelength, the S-loaded matrix absorbs less and as a result, no emission could be observed in the non-irradiated area. However, in the case of the irradiated area, both emission bands are still present, although red-shifted by 20 nm. As far as the PbS nanoparticles are concerned, the red-shift of the 650 nm emission band can be interpreted through a resonant size-selection by the excitation wavelength, as already observed with CdS particles [27]. The same kind of selection process in the electronic level population may be assumed for the "matrix band" at 650 nm. Meanwhile, the intensity ratio between these two emission bands is inverted when switching from UV to visible excitation. This can easily be understood as the result of a drastic decreasing number of absorbing entities in the non-irradiated zone between 351 and 514 nm wavelengths. The maximum of the emission band located around 930 nm is because of the presence of dominant small PbS nanoparticles (3-5 nm) [28]. This maximum emission is shifted toward lower wavelengths comparing to the one reported by Hines and Scholes [29] for a bigger colloidal PbS with a size of 6.5 nm.

In the literature, the mechanism of PbS nanoparticles formation inside a porous silica matrix has been explained by pyrolysis or decomposition of the PbS precursor under heat treatment [21,22]. Moreover, Chao et al. [24] have reported on the mechanism of PbS nanoparticles precipitation inside melting glasses induced by a femtosecond laser: when glasses containing lead and sulfur are submitted to a femtosecond laser irradiation, it can probably induce ion-exchange effects and a redistribution of network modifiers, resulting in the enrichment of the laser-irradiated zone in lead and sulfur atoms. This enrichment makes it possible to decrease the thermal treatment temperature. Hence, the formation of bigger and denser quantum dots in the irradiated area is accelerated after a subsequent and necessary heat treatment. In our case, the nano-crystallization of PbS inside the porous silica occurred in the region of the laser irradiation without any further annealing because of two main processes. First, the multi-photon absorption of the porous silica matrix and of the PbS precursors at 800 nm [30] leads to the decomposition of these precursors, thus enriching the irradiated zone with lead and sulfur ions. In the same time, the temperature elevation during the laser pulse could be sufficient to aggregate the molecular PbS into nanoparticles [26,31].

The effect of the incident average laser power on the size of the PbS nanocrystallites has been studied. Three S-doped samples were irradiated by the pulsed femtosecond laser at three different incident powers: 10, 25, and 40 mW. XRD measurements were performed on the irradiated samples, as shown in Figure 4. For each power, the XRD pattern presents reflections for 2θ equal to 26.1°, 29.9°, 42.9°, and 50.8°. These reflexes can be attributed to (111), (200), (220), and (311) reticular planes of the PbS cubic phase, respectively [21,22]. One can note an increase in intensity and a decrease in width of the reflexes with increasing the irradiation power. The mean nanocrystal diameter, d, can be determined from the line width of the XRD pattern by the Scherrer formula [32].
d = (0.94λ) / (B cos θ B )(1)
where λ is the X-ray wavelength, B is the full width at half maximum of the diffraction reflex (in radian), and θ B is the half angle position of the diffraction reflex on the 2θ scale. The diameter of the PbS nanoparticles, calculated from the Scherrer formula (Equation 1), is summarized in Table 1.

The most important drawback that limits the use of PbS nanoparticles in many applications is their low chemical stability. This is due to their high ability to oxidation, which limits their use under ambient conditions.

To indirectly investigate the stability of PbS nanoparticles precipitated inside the silica host matrix by pulsed laser irradiation, irradiated samples have been left in ambient air a for long period and optical absorption spectrum has been monitored upon time. Figure 5 shows the absorption spectra of two samples irradiated with 10 and 40 mW at three different times after the irradiation. One can note that the absorption edge of the sample irradiated with 40 mW was blue-shifted after 100 days. This blue-shift is attributed to the oxidation of PbS nanoparticles created inside the silica matrix. On the other hand, the absorption edge of the sample irradiated with 10 mW presents no shift after 100 days in comparison with the absorption taken just after preparation.

These preliminary results demonstrate that the stability of PbS nanoparticles, inside the porous host matrix, depends on their size. It should be recalled that the PbS particle mean size, as deduced from the XRD measurements around time t 0 , has been evaluated to 4 and 8 nm after irradiations with 10 and 40 mW, respectively (see Table 1). In the first case, the crystallite size is thus lower than the average matrix pore size of 5.4 nm, as opposed to the case of 40 mW irradiation. Since the diameter size of PbS nanoparticles created with 40 mW is greater than the pore diameter, it implies that, under the extreme local conditions of the femtosecond laserinduced plasma, the created nanoparticles are capable of breaking the walls of the interconnected pores during their formation. An important amount of oxygen is then allowed to diffuse inside the matrix and to react with PbS nanoparticles, leading to their oxidation in several days. It has been noticed that the oxidation rate seems to decrease versus time. This shift in the absorption band edge could be explained by the formation of coreshell structure with unknown shell phase. Indeed several oxide phases can be observed when PbS is oxidized such as PbO. PbSO 4 [33]. We have already detected this phase when we irradiate the doped monoliths by 514.5 nm continuous laser. On the contrary, no oxidation was observed with a lower laser power of 10 mW, but most of the nanoparticles have a size (4 nm) suited to the pores. In this case, it is likely that the nanoparticles have grown without any modification of the silica matrix and these particles remain protected from oxidation by the silica walls. This effect could explain the stability of PbS nanoparticles, even after 100 days. Hence, both the pore diameter of the host silica matrix and the power of pulsed femtosecond laser irradiation play an important role in the stability of the obtained PbS nanoparticles.

Conclusion

The formation of PbS nanoparticles localized inside porous silica xerogels has been performed using the pulsed femtosecond laser irradiation without any annealing. Characterization of the obtained nanocrystals was allowed by writing a tight network of PbS lines in a centimeter-sized zone near the surface or in the depth of the bulk xerogel. Most of the crystallites have been identified as cubic phase of PbS by XRD and TEM. We have shown that the power laser can be used to control the mean particle size between 4 and 8 nm. The micro-PL data made it possible to identify an emission band coming from the matrix and PbS precursors, and another band assigned to PbS nanoparticles, which are resonantly wavelength-selected in size. Finally, the stability of PbS nanoparticles under ambient conditions depends on the particle size, monitored by the laser power, and on the porosity of the host matrix.

Figure 1
1The irradiated area (10 mW) inside an S-doped silica monolith.

the TEM grid. Moreover, Figure 2c presents the HR-TEM of different nanoparticles. The calculation of the atomic interplanar distances has been performed on five zones labeled by circles. The zones 1, 2, and 3 exhibit fringe distances around 0.34 nm, which are attributed to the (111) lattice planes of cubic PbS (d 111 = 0.343 nm, JCPDS card, reference code 03-065-0692). While the fringes spacing for the zones 4 and 5 were of about 0.3 nm, which corresponds to the (200) lattice planes of PbS (d 200 = 0.297 nm, JCPDS card, reference code 03-065-0692). In addition, Figure 2d presents an electron diffraction pattern taken in a zone filled with nanoparticles. The estimated diffraction radii R1, R2, and R3 correspond well to the PbS lattice planes (200), (220), and (222), respectively.

Figure 2
2Results of TEM measurements in the irradiated area with 40 mW: (a) TEM image, (b) EDX spectrum obtained from an ensemble of PbS nanocrystals, (c) HR-TEM image of several PbS nanoparticles, (d) electron diffraction image taken from a zone filled with nanoparticles.

Figure 3
3Left: Optical absorbance spectra of S-doped silica matrix: (a) non-irradiated area, (b) femtosecond laser irradiated area with 40 mW. Right: (c) PL spectrum of a non-irradiated area under excitation at 351 nm, (d) PL spectrum of the irradiated area under excitation at 351 nm, (e) PL spectrum of a non-irradiated area under excitation at 514.5 nm and (f) PL spectrum of the irradiated area under excitation at 514.5 nm.

Figure 4
4XRD patterns of S-doped silica matrices recorded in the irradiated area with different laser powers: (a) 40 mW, (b) 25 mW and (c) 10 mW.

Figure 5
5Optical absorption of PbS nanoparticles created by femtosecond laser at different times. Left: sample irradiated with 40 mW, Right: sample irradiated with 10 mW. (a) 100 days after the initial irradiation, (b) 50 days after the initial irradiation and (c) just after the initial irradiation.

Table 1
1Calculated sizes of PbS nanoparticles from XRD measurements for different laser average powersPower (mW)Sizes calculated from XRD (nm)
AcknowledgementsThis study was supported by the French Agence Nationale de la Recherche (ANR) in the frame of the POMESCO project (Organized Photo-growth of Metallic and Semi-Conductor Nano-Objects Intended to Optic Devices), the "Conseil Régional Nord Pas de Calais Picardie" and the "Fonds Européen de Développement Economique des Régions".Authors' contributions AC and HE carried out the preparation of the samples as well as the writing of the manuscript. RB and LB have preformed the XRD analysis and the calculation of sizes. LB carried out the PL analysis. MLP is responsible for the femtosecond laser operations. AC, HE, OC, BC, and MB have interpreted the results, as well as drafted the manuscript. All authors read and approved the final manuscript.Competing interestsThe authors declare that they have no competing interests.
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Direct-writing of PbS nanoparticles inside transparent porous silica monoliths using pulsed femtosecond laser irradiation. Chahadih, Convenient online submission 7 Rigorous peer review 7 Immediate publication on acceptance 7 Open access: articles freely available online 7 High visibility within the fi eld 7. 6542Cite this article as. Retaining the copyright to your article Submit your next manuscript at 7 springeropen.comCite this article as: Chahadih et al.: Direct-writing of PbS nanoparticles inside transparent porous silica monoliths using pulsed femtosecond laser irradiation. Nanoscale Research Letters 2011 6:542. Submit your manuscript to a journal and benefi t from: 7 Convenient online submission 7 Rigorous peer review 7 Immediate publication on acceptance 7 Open access: articles freely available online 7 High visibility within the fi eld 7 Retaining the copyright to your article Submit your next manuscript at 7 springeropen.com

. Chahadih, Nanoscale Research Letters. 6542Chahadih et al. Nanoscale Research Letters 2011, 6:542


Case report: a unique pediatric case of a primary CD8 expressing ALK-1 positive anaplastic large cell lymphoma of skeletal muscle

Timo Gaiser
Eva Geissinger
Torsten Schattenberg
Hanns-Peter Scharf
Matthias Dürken
Dietmar Dinter
Andreas Rosenwald
Alexander Marx
Case report: a unique pediatric case of a primary CD8 expressing ALK-1 positive anaplastic large cell lymphoma of skeletal muscle
C A S E R E P O R T Open AccessALK-1Anaplastic large cell lymphomaCD30Pediatric lymphoma
Primary involvement of skeletal muscle is a very rare event in ALK-1 positive anaplastic large cell lymphoma (ALCL). We describe a case of a 10-year old boy presenting with a three week history of pain and a palpable firm swelling at the dorsal aspect of the left thigh. Histological examination of the lesion revealed a tumoral and diffuse polymorphic infiltration of the muscle by large lymphoid cells. Tumor cells displayed eccentric, lobulated "horse shoe" or "kidney-shape" nuclei. The cells showed immunohistochemical positivity for CD30, ALK-1, CD2, CD3, CD7, CD8, and Perforin. Fluorescence in situ hybridization analysis revealed a characteristic rearrangement of the ALK-1 gene in 2p23 leading to the diagnosis of ALK-1 positive ALCL. Chemotherapy according to the ALCL-99-NHL-BFM protocol was initiated and resulted in a complete remission after two cycles. This case illustrates the unusual presentation of a pediatric ALCL in soft tissue with a good response to chemotherapy.

Background

Anaplastic large cell lymphoma (ALCL) was discovered by Stein et al. and characterized by strong expression of the Ki-1 (CD30) antigen [1]. In 1988, ALCL was included in the revised Kiel classification and is nowadays classified as a Non-Hodgkin lymphoma of T-cell origin by the World Health Organization [2,3]. The key moment in understanding the primary biological driver behind ALCL was the discovery of the recurrent t(2;5)(p23;q35) translocation in this lymphoma type [4]. This finding was further elucidated by Morris and colleagues who identified the two genes involved in this translocation: the anaplastic lymphoma kinase (ALK-1) on chromosome 2 and nucleophosmin (NPM1) on chromosome 5 [5]. Subsequently, the NPM-ALK fusion protein was proven to have oncogenic capacity [6]. Among the four main types of pediatric non-Hodgkin lymphoma (Burkitt lymphoma, diffuse large Bcell lymphoma, anaplastic large cell lymphoma and lymphoblastic lymphoma) ALCL has the best prognosis: event-free and overall survival rates were 72% and 88%, respectively, even in advanced-stage diseases [7].

Here, we report a case of a 10-year old boy diagnosed with an ALK-1 positive anaplastic large cell lymphoma of the skeletal muscle. Primary involvement of skeletal muscle by ALCL is extremely rare and so far only four pediatric cases have been reported in the literature [8][9][10][11].

Case presentation

Initial presentation and management A 10-year old boy presented with a painful swelling in the dorsal aspect of the left distal thigh without history of trauma. Pain started 3 weeks before, increased with time and resulted in difficulties in walking at time of presentation. No fever, night sweats or weight loss was reported. Physical examination revealed a 9 × 4 cm palpable firm mass in the biceps femoris muscle. Magnetic resonance imaging (MRI) demonstrated a 4 × 3 × 9 cm ill-defining tumor within the long head of the biceps femoris muscle with vivid uptake of contrast medium ( Figure 1). Also, enlarged lymph nodes up to a diameter of 2 cm were detected popliteal, inguinal and iliac. Radiologically, the most probable diagnosis was a malignant soft tissue tumor, most likely a rhabdomyosarcoma. Beside slightly enlarged inguinal and popliteal lymph nodes further staging did not reveal other organ or skin involvement. Laboratory HIV 1 + 2 screening tests showed no signs for HIV infection. Histological diagnosis was achieved by incisional biopsy. After being classified as Murphy stage II [12], further treatment was conducted according to the international protocol for childhood ALCL (ALCL-99-NHL-BFM). ALK-1/NPM PCR was found positive in peripheral blood. After two cycles of chemotherapy the tumor was no longer detectable clinically or by ultrasound. The patient is currently receiving further chemotherapy without experiencing major toxicity.

Materials and methods

Immunohistochemistry

In situ hybridization

In situ hybridization analyses for the t(2;5) translocation and for Epstein-Barr virus (EBV) encoded small nuclear RNAs (EBERs) was performed on four micron sections of formalin fixed, paraffin-embedded tissue paraffin sections using the ALK Dual Color, Break Apart Rearrangement Probe (Abbott, North Chicago, IL, USA) and ZytoFast™ EBV Probe (ZytoVision GmbH, Bremerhaven, Germany); procedures were performed according to the manufacturers' indications.

Results

Histologically, the muscle of the biceps femoris displayed a diffuse polymorphic lymphoid infiltration of high mitotic activity, including atypical mitoses. While no haemangio invasion was demonstrable, lymphovascular invasion was detectable morphologically and by D2-40 immunohistochemistry. The majority of tumor cells was small to middle sized with pleomorphic nuclei and variable amount of pale cytoplasm. Intermingled and less common were larger cells with eccentric lobulated "horse shoe" or "kidneyshape" nuclei and abundant cytoplasm with an intense eosinophilic region representing the Golgi apparatus (Figure 2A). These "hallmark cells" strongly expressed CD30, while the smaller lymphoid cells were only moderately positive ( Figure 2B). Furthermore, the tumor cells stained positive for CD2, CD3, CD7 ( Figure 2C), CD99, Perforin and, remarkably, CD8 ( Figure 2D). ALK-1 had an obvious cytoplasmic and nuclear pattern of immunoreactivity in the tumor cells ( Figure 2E). As expected, rearrangement of the ALK-1 gene in 2p23 could be proven by fluorescence in situ hybridization analyses ( Figure 2F). The whole tumor cell population revealed a CD5 antigen loss and a weak and obviously only cytoplasmic reactivity for βF1. The Ki67 proliferation index reached~60%. Tumor cells stained negative for CD10, CD20, CD79a, CD34, CD56, CD117, TdT, Desmin, and EMA. With EBER in situ hybridization no EBV positive cells were detectable. Based on these findings the diagnosis of CD30/ALK-1 positive anaplastic large cell lymphoma was established and confirmed by the reference center for lymphoid malignancies in Würzburg (AR, EG).

Conclusions

ALCL can be classified into ALK-1 negative and ALK-1 positive ALCL. ALK-1 negative ALCL occurs mainly in older patients (peak incidence~60 y) and in advancedstage disease. ALK-1 positive ALCL is a clinically aggressive lymphoma that mostly occurs in the first three decades of life [13]. However, overall survival and longer disease free survival is observed after treatment with aggressive chemotherapy. Patients affected by ALK-1 positive ALCL have a significant better overall survival than ALK-1 negative ALCL patients (5-year overall survival: 70-80% vs. 33-49%) [14,15]. It is still controversially discussed if this observation can be explained by the biological role of the ALK-1 fusion proteins or by the younger age of the ALK-1 positive ALCL patients. Unlike ALK-1 positive ALCL, which is characterized by t(2;5)(p23;q35) translocation and resulting in the expression of the NPM-ALK-1 fusion protein, so far no recurrent cytogenetic alterations have been described in ALK-1 negative ALCL.

The presented case illustrates a CD30+, ALK-1+ ALCL of T-cell origin (CD2-/+, CD3+, CD7+, CD8+ and Perforin+) in a pediatric patient. Due to the abundance of small neoplastic cells with pale cytoplasm mixed with some medium-sized and large lymphoid cells this case is best counted among the "small cell variant" of ALCL (10%). Other ALCL subcategories are the common (70-80%) and lymphohistiocytic type (10%).

Primary malignant lymphoma of soft tissue is an infrequent, and often diagnostically challenging neoplasm [16,17]. Among those studies which reported the T-or B-cell phenotype of primary soft tissue lymphoma, B-cell accounted for over 90%. Most of those cases showed aggressive histology and were of diffuse large B-cell phenotype [17]. T cell lymphomas in general and ALCL in FISH with ALK Dual Color, Break Apart Rearrangement Probe for detecting translocations affecting 2p23 ALK breakpoint region. ALK locus in its native state is seen as two immediately adjacent red and green signals. However, if a t(2;5) chromosome rearrangement has occurred as in this case, one separate red and one separate green signals can be seen (the one remaining native ALK remains as an orange/green signal) (magnification 1000×). particular exceedingly rarely exhibit primary infiltration of skeletal muscle [8][9][10]. Of note, all intramuscular primary ALCL cases reported so far have been ALK-1 positive. Why this T-cell lymphoma could overcome the normally well established protection of skeletal muscle against lymphocytic infiltration is unknown [18].

Interestingly, the current ALCL is the first among the reported intra-muscular ALCL primaries to show a CD8 phenotype, which is extremely rare in ALCL. However, trauma, which theoretically could have acted as a stimulatory trigger providing signals for T cell homing to skeletal muscle [18] was denied by the patient. Moreover, no other infectious or autoimmune process was known from the history or the local findings.

It should also be mentioned in this context that the slightly enlarged lymph nodes were only detected after MRI examination and were not clinically indicative. Unfortunately a lymph node biopsy was not performed. Therefore, a final statement about a possible secondary involvement of the muscle was not possible. However, we consider this possibility less likely because the majority of ALCL patients (70%) presents with advanced stage III to IV disease [3], which was obviously not the case in our patient.

ALCL has to be distinguished from classical Hodgkin lymphoma (cHL), CD30+ non-Hodgkin B cell lymphomas and very rare ALK-1 positive (and eventually CD30negative) large B cell lymphomas. Differential diagnosis between ALCL and cHL can be made by expression of cytotoxic molecules such as Granzyme B, Perforin and T-cell-restricted intracellular antigen-1 (TIA1), EMA and CD45/LCA which are typical of ALCL, while positivity for CD15 (in 70% of cases), PAX5 (90%) and LMP1 (in 50%) are typical of cHL. To distinguish ALCL from ALK-1-positive large B cell lymphomas the lack of CD30 expression is critical and the t(2;5) translocation cannot be demonstrated in the B cell lymphoma [19].

We conclude that this case represents a very rare manifestation of an ALCL in soft tissue. Despite the anaplastic appearance a favorable outcome was possible after promptly applying aggressive chemotherapy. The expression of the NMP-ALK-1 protein and the young age of the patient might have influenced the positive outcome.

Consent

Written informed consent was obtained from the patient, respectively the guardians for publication of this case report and accompanying images. A copy of the written consent is available for review to the Editor-in-Chief of this journal.

Figure 1
1Axial MRI of the left thigh delineates an ill-defining tumor mass within the long head of the biceps femoris muscle with vivid uptake of contrast media while measuring 4 × 3 × 9 cm in size.

Figure 2
2Histological features and immunophenotype of the intramuscular ALCL. (A) The biceps femoris muscle showed a tumoral and diffuse polymorphic infiltration by large lymphoid cells (magnification 100×). Inlay: Majority of the cells were small to middle sized with pleomorphic nuclei and various size of pale cytoplasm. Intermingled and less common larger cells with eccentric lobulated "horse shoe" or "kidney-shape" nuclei and an abundant cytoplasm (so called "hallmark cells") (magnification 400×). (B) Strong membranous and perinuclear dotlike positivity for CD30 (magnification 100×). (C) T-cell phenotype of ALCL demonstrated by positivity for CD7 and CD8 (D) (magnification 100×). (E) Tumor cells expressing ALK-1 protein both in the cytoplasm and nucleus indicating the classical t(2;5) translocation (magnification 200×). (F)
© 2012 Gaiser et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Gaiser et al. Diagnostic Pathology 2012, 7:38 http://www.diagnosticpathology.org/content/7/1/38
Authors' contributions TG and AM drafted the manuscript and analysed the data. EG and AR served as reference pathologists. TS, HPS and MD were leadingly involved into the clinical examination and the treatment of the patient. DD carried out MRI and subsequent imaging analyses. All authors read and approved the manuscript.Competing interestsThe authors declare that they have no competing interests.
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Submit your next manuscript to BioMed Central and take full advantage of: • Convenient online submission • Thorough peer review • No space constraints or color figure charges • Immediate publication on acceptance • Inclusion in PubMed, CAS, Scopus and Google Scholar • Research which is freely available for redistribution. Gaiser, Diagnostic Pathology. 738Case report: a unique pediatric case of a primary CD8 expressing ALK-1 positive anaplastic large cell lymphoma of skeletal muscle. Submit your manuscript at www.biomedcentral.com/submitCite this article as: Gaiser et al.: Case report: a unique pediatric case of a primary CD8 expressing ALK-1 positive anaplastic large cell lymphoma of skeletal muscle. Diagnostic Pathology 2012 7:38. Submit your next manuscript to BioMed Central and take full advantage of: • Convenient online submission • Thorough peer review • No space constraints or color figure charges • Immediate publication on acceptance • Inclusion in PubMed, CAS, Scopus and Google Scholar • Research which is freely available for redistribution Submit your manuscript at www.biomedcentral.com/submit