Dear Editor

We appreciated the comments of Dr Zardawi [1] and agree that actin is a ubiquitous cytoskeletal protein of microfilaments and demonstrable in a variety of cells and tumor types. As we described, all leiomyosarcomas are smooth muscle actin (SMA)-positive, and desmin, muscle specific actin (MSA) and h-caldesmon are positive in a great majority of these tumors. However, none of these is absolutely specific for smooth muscle, and positivity for two or more of these markers is more supportive of leiomyosarcoma than positivity for one alone.

Anti-SMA monoclonal antibody, 1A4, detects mainly with an alpha smooth muscle actin isoform. This marker exhibits a more restricted pattern of staining for smooth muscle, but this may be expressed in rhabdomyosarcomas and in cells with a myofibroblastic or myoepithelial phenotype. As another anti-MSA monoclonal antibody, HHF35, recognizes all alpha actins (skeletal, smooth, and cardiac) and gamma smooth muscle actin, its specificity for smooth muscle is therefore quite limited. In this regard, recognition of the fact that non-muscle lesions exhibiting so-called myoid differentiation are also SMA-positive will prevent overdiagnosis as leiomyosarcoma.

Reference

(1) Zardawi IM. Re: Expression of smooth muscle markers in so called malignant fibrous histiocytoma [electronic response to Hasegawa et al Expression of smooth muscle markers in so called malignant fibrous histiocytomas] jclinpath.com 2003http://jcp.bmjjournals.com/cgi/eletters/56/9/666#45

Dear Editor

I read with interest the article on the expression of smooth muscle markers in so called malignant fibrous histiocytomas by Hasegawa et al. in the Journal.[1]

While I agree with Hasegawa and colleagues that pleomorphic malignant fibrous histiocytoma can be regarded as an undifferentiated sarcoma, I feel Hasegawa et al are placing too much reliance on so called smooth muscle markers. Most of these markers are ubiquitous, present in many cells. Many of the so called muscle specific markers stain microfilaments which form the cytoskeletal framework of most cells. However, these markers are particularly expressed in muscle cells, myoepithelial cells cell and myofibroblasts. In neoplasia, these markers indicate myogenic differentiation which can be seen in a raft of tumours which make them unreliable in tumour classification.

A search of the Internet (www.ipox.org) shows smooth muscle actin (SMA) immunoreactivity in 150 types of tumour from different parts of the body with 100% staining in 32 tumour types, between 50% and 99% staining in 45 tumour types, and between 4% and 49% staining in 73 tumour types. The same source cites 0% staining in 80 tumour types. The question that needs to be asked therefore is not which tumours are SMA positive but rather which tumours do not express this marker.

SMA is very much like neuron specific enolase (NSE) which can be found in neurons, neuroendocrine cells, striated and smooth muscle cells, megakaryocytes, T cells and platelets to name a few. NSE reactivity has been reported in over 140 tumour types (www.ipox.org).

Reference

(1) Hasegawa T, Hasegawa F, Hirose T, Sano T, Mastuno Y. Expression of smooth muscle markers in so called malignant fibrous histiocytoma. J Clin Pathol 2003; 56:666-671.

Dear Editor

We read the article by Horny et al. describing bone marrow mast cell (MC) specific protease expression patterns in cases of systemic mastocytosis and myelodysplastic syndromes (MDS) with great interest.[1] Increase in bone marrow MC is a known feature of various hematological diseases including myeloproliferative disorders and acquired severe aplastic anemia (SAA). Although the MC increase is clonal in mastocytosis and benign in acquired SAA, its nature has not been fully understood in myeloproliferative and myelodysplastic disorders.

Acquired SAA and hypoplastic MDS share several clinical and bone marrow features and often difficult to distinguish. Both conditions respond to immune suppressive therapies. Is the increased MC in these conditions simply, an innocent consequence of hemopoietic cell injury sparing MC or alternatively, does it contribute to the development of severe bone marrow hypoplasia/aplasia in return? Mast cells have long life spans and it is likely that they were not directly affected by the attack against stem cell compartment resulting in relative MC increase in the bone marrow. Low to normal stem cell factor (SCF) levels has been shown in SAA unlike the increased levels of other hemopoietic growth factors.[2] This may be explained by greater dependency of MC survival and growth on SCF than other growth factors and by a negative feedback control mechanism in a population that is already supplied by an autocrine pathway. In support of this explanation, a reaction mimicking systemic mastocytosis was observed in an aplastic anemia patient treated with SCF accompanied by a partial and transient hemopoietic recovery.[3]

Mast cells with various enzyme expression patterns may mediate different functions in certain tissues that they exist in. These patterns may also be related to the maturational stage of MC. Nevertheless, the predominant MC type in certain tissues may be determined by the environmental needs. We think that coexistence of chymase-expressing MC (MCC) and chymase and tryptase-expressing MC (MCCT) in physiological conditions reflects naturally occurring balance contributing to tissue homeostasis. It is known that MC can also act as antigen presenters as well as effector elements of human immune system. Mast cells can kill target cells through secretion of cytokines such as TNF-a and serine proteases and potentially through direct cell-to-cell interaction. Granzyme H, one of the MC serine proteases, has chymase activity [4] and chymase is known to induce apoptosis in target cells.[5] It was also demonstrated that mast cell-derived cell line P815 contains granzyme B RNA.[6] On the other hand, tryptase, another MC protease, is a well-known mitogen that could induce growth of certain cells such as airway smooth muscle cells, fibroblasts, neuronal cells.[7] Tryptase-expressing MC (MCT) are often found in tissue repair sites characterized by fibrosis.

The predominance of MCT in systemic mastocytosis and MDS patients was consistent with the typical presence of hypercellular marrow in these conditions.[1] Although authors did not provide the number of cases with hypoplastic MDS in their series, the frequency has been 5-10% in adult literature suggesting that the majority, if not all of their cases had normocellular or hyperplastic MDS. The autocrine production of SCF with increased tryptase activity might have contributed to the extremely hypercellular bone marrow in those cases. The authors also described hypocellular bone marrow associated with a focal increase in MC with strong chymase expression in a case of indolent systemic mastocytosis that suggests a possible MCC contribution to hypocellularity. We recently showed an association between MC persistence and poor outcome in childhood SAA following immune suppression.[8] In another study, we demonstrated long- term liquid culture-grown human bone marrow MC cytotoxicity against human leukemia cells.[9] It is possible that those MC had strong chymase expression. Regardless of the mechanisms involved, MC, preferentially MCC increase may contribute to hypocellularity in acquired SAA and hypoplastic MDS. This explanation is also consistent with the lack of fibrosis in acquired SAA and hypoplastic MDS that could be secondary to specific MCC increase.

References

(1) Horny H-P, Greschniok A, Jordan J-H, Menke DM, Valent P : Chymase expressing bone marrow mast cells in mastocytosis and myelodysplastic syndromes: an immunohistochemical and morphometric study. J Clin Pathol 2003;56:103-106.

(2) Kojima S, Matsuyama T, Kodera Y. Plasma levels and production of soluble stem cell factor by marrow stromal cells in patients with aplastic anaemia. Br J Haematol 1997;99:440-6.

(3) Jordan JH, Schernthaner GH, Fritsche-Polanz R, et al. Stem cell factor- induced bone marrow mast cell hyperplasia mimicking systemic mastocytosis (SM): histopathologic and morphologic evaluation with special reference to recently established SM-criteria. Leuk Lymphoma 2002;43:575-82.

(4) Edwards KM, Kam CM, Powers JC, Trapani JA. The human cytotoxic T cell granule serine protease granzyme H has chymotrypsin-like (chymase) activity and is taken up into cytoplasmic vesicles reminiscent of granzyme B-containing endosomes. J Biol Chem 1999;274:30468-73.

(5) Leskinen M, Wang Y, Leszczynski D, Lindstedt KA, Kovanen PT. Mast cell chymase induces apoptosis of vascular smooth muscle cells. Arterioscler Thromb Vasc Biol 2001;21:516-22.

(6) Garcia-Sanz JA, MacDonald HR, Jenne DE et al. Cell specificity of granzyme gene expression. J Immunol 1990;145:3111-8.

(7) Ruoss SJ, Hartmann T, Caughey GH. Mast cell tryptase is a mitogen for cultured fibroblasts. J Clin Invest 1991;88:493-9.

(8) Chien M, Abella E, Rabah R, Ravindranath Y, Savasan S. Mast cell persistence is associated with poor outcome in childhood severe aplastic anemia following immune suppression. Presented in the 17th Annual Meeting of ASPH/O, Seattle, Washington, May 3- May 6, 2003. Pediatr Res 2003;53:292A, Abstract #1663.

(9) Özdemir Ö, Ravindranath Y, Savasan S. Evaluation of long-term liquid culture grown human bone marrow mast cell cytotoxicity against human leukemia cells. (44th annual meeting of the American Society of Hematology, Philadelphia, Pennsylvania, December 6-10, 2002.) Blood 2002; 100:45b, Abstract #3642.

Dear Editor

The article entitled “Health and Safety at necropsy” by Julian Burton provides a detailed and well written narrative regarding both the risks and hazards faced by professionals during post-mortem examinations.[1]

Despite the presence of a relatively large publication base regarding this topic, important aspects are highlighted, including transmissible spongiform encephalopathies and the more modern, but potentially dangerous, advances in medical technologies. However, we would wish to clarify the issues the author raises regarding exploding bullets. The difference between a true exploding bullet and a projectile designed to fragment on impact is one of great importance, and one that may cause confusion, as would appear to be the case within this article.

Bullets are composed of a casing containing an explosive powder charge which, on striking, forces the end projectile element out at speeds of up to 1500 metres per second, depending upon the ammunition and the type of gun used. The projectile causes soft tissue damage through crushing, creating a temporary cavity which contains hot gases. The tissue is compressed radially from the centre of the cavity and, depending on its elastic properties, results in tears to structures (as seen with injuries to solid abdominal viscera). The recoil of the tissues, together with the dissipation of the gases, causes the soft tissue to collapse inwards on itself, the resultant defect being the permanent cavity.

Expansion, or hollow-point, bullets are specialised bullets designed to deform upon impact due to a collapsible space within the projectile tip. The result is that a single projectile will inflict greater overall damage to a target, allowing an increased transfer of kinetic energy compared to a standard bullet. The “benefits” include a decreased risk of ricochet as the overall penetration distance is reduced, however, some of the older ammunition failed to expand on impact due to pieces of clothing obstructing the cavity.

Pre-fragmented, or frangible, bullets are composed of a pre-scored outer jacket with a plastic round nose containing compressed lead shot within. The result is a controlled explosion on impact producing increased damage and less clothing related problems. The tips, however, possess no explosive charge.

Burton describes the Winchester Black Talon SXT bullet, though erroneously includes this within the heading of exploding bullets.[1] It is, in fact, a type of expansion bullet. The tip is coated with a black lubricant and has a hollow-point possessing six pre-scored serrations designed to rapidly open outwards upon impact. The jacket of the bullet is thickest at its tip, unlike most hollow-point bullets, to provide support for the claw-like petals as the bullet passes through the body. The result, in theory, is a wider permanent cavity created by a single projectile thus increasing the likelihood of damage to a vital structure. The bullet was voluntarily removed from the market in 1994 and remarketed as the Ranger SXT, and later as the Ranger Talon, both available only to law enforcement officers. Despite media assertions, these projectiles are not “armour-piercing”, the title relating purely to a widely reported manufacturing error in one brand of body armour, which resulted in a recall of this product. Although such expansion bullets do indeed pose a health and safety hazard, due to the sharp edges of the deformed projectile, there is no risk of explosion at necropsy.

True exploding bullets were first described over a century ago, and, though not actually in use at that time, were prohibited under the St. Petersburg Declaration of 1868, which states that explosive or inflammable projectiles, with a weight of less than 400 grams, should never be used in the time of war. Examples include the Russian 7.62mm x 54R machine gun ammunition with an internal charge of tetryl and phosphorus, and later handgun cartridges containing Pyrodex charges with or without mercury additives.[2] It should also be noted that individuals can easily obtain instructions for the creation of their own bullets. The most infamous use of such bullets was the attempted assassination of President Reagan in 1981 by John Hinckley, who used “Devastator” bullets (Bingham Limited, USA) composed of a lacquer-sealed aluminium tip with a lead azide centre designed to explode on impact. Though frequently referred to in works of fiction, they are rarely encountered in forensic practice, as sales have been restricted following the incident in 1981. Projectiles that have failed to detonate are also not as sensitive to movement and heat as mentioned in the article; the author refers to an article on this topic, but fails to acknowledge a follow-up letter correcting Knight’s original mistakes. [2,3] Burton has, unfortunately, reproduced these errors in his text. Additionally, unexploded bullets are safe on exposure to X-rays and ultrasound.[4] The quantity of explosive is small and, if it fails to detonate on high-velocity impact, is unlikely to explode during autopsy examination. We would indeed agree with the assertion that safety glasses be used during necropsy examination of ballistic victims, however, as Burton himself details within his own book, such eye protection should be routine practice regardless of the cause of death.[5]

A footnote on the topic would include the use of armour-piercing incendiary round ammunition employed during recent conflicts that possess explosive points, such as the Raufoss Multipurpose Projectiles, (Nammo, Norway), fired from anti-vehicle guns of varying calibre.[6] These are not designed, nor are they produced for use against personnel. In fact, the round will pass through the body unexploded and are thus unlikely to be present in bodies from military conflicts. As such, it is also argued that they do not contravene the St. Petersburg Declaration. If present with a body, they are safe to handle, transport and store. They also comply with NATO standards ensuring complete handling safety, even following vertical drops of up to 15 metres.

Finally, it is interesting to note that the Devastator bullet was developed in the 1970s for use by sky marshals, to minimise the risk of penetration of the plane fuselage when incapacitating a hijacker; a concept that appears to be return in light of recent world events.

References

(1) Burton, J.L. Health and Safety at necropsy. J Clin Pathol 2003;56(4):254-60.

(2) Conway, G.D., Jacobs, A. Explosive bullets: a new hazard for doctors. BMJ 1982;284:1707.

(3) Knight, B. Explosive bullets: a new hazard for doctors. BMJ 1982;284:768-9.

(4) Schlager, D., Johnson, T., McFall, R. Safety of imaging exploding bullets with ultrasound. Ann Emerg Med 1996;289(20):183-7.

(5) Burton, J.L., Rutty, G.N. The Hospital Autopsy, Second Edition. London: Arnold Publishing, 2001

(6) Nordic Ammunition Company website http://www.nammo.com