Epithelioid Fibrous Histiocytoma With Chondroblastoma-Like Features: A Report of a Rare Entity and Discussion of Related Diagnostic Challenges.

ABSTRACT: Epithelioid fibrous histiocytoma (EFH) is an uncommon benign skin lesion. It is distinct from FH by virtue of its recurrent anaplastic lymphoma kinase (ALK) gene rearrangements and immunohistochemical expression of ALK protein. It often poses a challenge in interpretation. Clinically, it is characterized by a flesh-colored papule/nodule on an extremity of a young to middle-aged individual. Microscopically, it is represented by a circumscribed dermal papule/nodule composed of sheets of plump epithelioid cells, forming whorled aggregates around numerous intralesional vessels. Immunohistochemistry, notably ALK positivity and relevant negative stains, serves to distinguish EFH from its morphological mimics. Rare examples of chondroblastoma-like EFH and EFH with osseous metaplasia are recorded in the literature. Our case is of a 58-year-old man who attended an oculoplastic surgeon because of an exophytic cutaneous nodule on the right upper eyelid. The lesion was excised. Microscopically, it displayed morphological and immunohistochemical features of EFH. Of interest, discrete foci of chondro-osseous change, including chondroblastoma-like pericellular calcification, osteoid formation, and osteoclast-like giant cells, were noted throughout the lesion. A diagnosis of EFH with chondroblastoma-like features was made. Of interest, the changes observed in this EFH serve to link the previously reported examples of pure chondroblastoma-like EFH and EFH with osseous metaplasia. This morphological variant of EFH adds to the existing diagnostic challenge presented by these lesions, particularly in the distinction from other calcifying tumors of the skin.

Recurrent IgG4-related tubulointerstitial nephritis concurrent with chronic active antibody mediated rejection: A case report.

IgG4-related disease is a relatively newly described entity that can affect nearly any organ, including the kidneys, where it usually manifests as tubulointerstitial nephritis (IgG4-TIN). The diagnosis can be suggested by characteristic histological features, including an inflammatory infiltrate with increased IgG4-positive plasma cells associated with "storiform" fibrosis. Serum IgG4 is usually elevated. In the native kidney and other organs, there is typically a brisk response to treatment with immunosuppression. Recurrence of IgG4-TIN after renal transplant has not been described in the literature. Here, we describe the first case of recurrent IgG4-TIN in a young patient concomitant with chronic active antibody mediated rejection five years after kidney transplant. Recurrent IgG4-TIN could be diagnosed by the characteristic histopathologic features and increased IgG4-positive plasma cells. Despite maintenance immunosuppression, this disease may recur in the kidney allograft.

In vitro cultivation of Hematodinium sp. isolated from Atlantic snow crab, Chionoecetes opilio: partial characterization of late developmental stages.

Hematodinium is a parasitic dinoflagellate of numerous crustacean species, including the economically important Atlantic snow crab, Chionoecetes opilio. The parasite was cultured in vitro in modified Nephrops medium at 0 °C and a partial characterization of the life stages was accomplished using light and transmission electron microscopy (TEM). In haemolymph from heavily infected snow crabs two life stages were detected; amoeboid trophonts and sporonts. During in vitro cultivation, several Hematodinium sp. life stages were observed: trophonts, clump colonies, sporonts, arachnoid sporonts, sporoblasts and dinospores. Cultures initiated with sporonts progressed to motile dinospores; however, those initiated with amoeboid trophonts proliferated, but did not progress or formed schizont-like stages which were senescent artefacts. Plasmodial stages were associated with both trophonts and sporonts and could be differentiated by the presence of trichocysts on TEM. Macrodinospores were observed but not microdinospores; likely due to the low number of Hematodinium sp. cultures that progressed to the dinospore stage. No early life stages including motile filamentous trophonts or gorgonlocks were observed as previously noted in Hematodinium spp. from other crustacean hosts. All Hematodinium sp. life stages contained autofluorescent, membrane-bound electron dense granules that appeared to degranulate or be expelled from the cell during in vitro cultivation.

The effects of temperature and body size on immunological development and responsiveness in juvenile shortnose sturgeon (Acipenser brevirostrum).

Sturgeon are an important evolutionary taxa of which little is known regarding their responses to environmental factors. Water temperature strongly influences growth in fish; however, its effect on sturgeon immune responses is unknown. The objective of this study was to assess how 2 different temperatures affect immune responses in shortnose sturgeon (Acipenser brevirostrum) relevant immune organs such as the meningeal myeloid tissue, spleen, thymus and skin. These responses were studied in 2 different sizes of same age juvenile sturgeon kept at either 11 °C or 20 °C (4 treatment groups), before and after exposure to an ectoparasitic copepod (Dichelesthium oblongum). Based on a differential cell count, temperature was found to strongly influence immune cell production in the meningeal myeloid tissue, regardless of the fish sizes considered. Morphometric analysis of splenic white pulp showed a transient response to temperature. There were no differences between the groups in the morphometric analysis of thymus size. Splenic IRF-1 and IRF-2 had similar expression profiles, significantly higher in fish kept at 20 °C for the first 6 weeks of the study but not by 14 weeks. In the skin, IRF-1 was significantly higher in the fish kept at 11 °C over the first 6 weeks of the study. IRF-2 had a similar profile but there were no differences between the groups by the end of the trial. In conclusion, higher water temperatures (up to 20 °C) may have beneficial effects in maximizing growth and improving immunological capacity, regardless of the fish sizes considered in this study.

The transcription factor Sry-related HMG box-4 (SOX4) is required for normal renal development in vivo.

BACKGROUND: The DNA-binding transcription factor Wilms' Tumor Suppressor-1 (WT1) plays an essential role in nephron progenitor differentiation during renal development. We previously used Wt1 chromatin-immunoprecipitation coupled to microarray (ChIP-chip) to identify novel Wt1 target genes that may regulate nephrogenesis in vivo. We discovered that all three members of the SoxC subfamily, namely, Sox4, Sox11, and Sox12, are bound by Wt1 in mouse embryonic kidneys in vivo. SoxC genes play master roles in determining neuronal and mesenchymal progenitor cell fate in a multitude of developmental processes, but their function in the developing kidney is largely unknown. RESULTS: Here we show that all three SoxC genes are expressed in the nephrogenic lineages during renal development. Conditional ablation of Sox4 in nephron progenitors and their cellular descendants (Sox4(nephron-) mice) results in a significant reduction in nephron endowment. By postnatal day (P)7, Sox4(nephron-) renal corpuscles exhibit reduced numbers of Wt1+ podocytes together with loss of expression of the slit diaphragm protein nephrin. Sox4(nephron-) mice develop early-onset proteinacious glomerular injury within 2 weeks of birth progressing to end-stage renal failure within 5-9 months. CONCLUSIONS: Collectively, our results demonstrate an essential requirement of Sox4 for normal renal development in vivo.

Osteoprogenitor cell therapy in an equine fracture model.

OBJECTIVE: To compare the efficacy of osteoprogenitors in fibrin glue to fibrin glue alone in bone healing of surgically induced ostectomies of the fourth metacarpal bones in an equine model. STUDY DESIGN: Experimental. ANIMALS: Adult horses (n = 10). METHODS: Segmental ostectomies of the 4th metacarpal bone (MC4) were performed bilaterally in 10 horses. There was 1 treatment and 1 control limb in each horse. Bone defects were randomly injected with either fibrin glue and osteoprogenitor cells or fibrin glue alone. Radiography was performed every week until the study endpoint at 12 weeks. After euthanasia, bone healing was evaluated using radiography and histology. Analysis of radiographic data was conducted using a linear-mixed model. Analysis of histologic data was conducted using a general linear model. Statistical significance was set at P < .05. RESULTS: Radiographic grayscale data as a measure of bone healing revealed no significant difference between treatment and control limbs. Radiographic scoring results also showed that the treatment effect was not significant. Histologic analysis was consistent with radiographic analysis showing no significant difference between the area of bone present in treatment and control limbs. CONCLUSION: Injection of periosteal-derived osteoprogenitors in a fibrin glue carrier into surgically created ostectomies of MC4 does not accelerate bone healing when compared with fibrin glue alone.

Ontogeny and disease responses of Langerhans-like cells in lymphoid tissues of salmonid fish.

The ontogeny and disease responses of Langerhans-like cells within lymphoid tissues of Atlantic salmon, Salmo salar, and rainbow trout, Oncorhynchus mykiss, were investigated. These cells were studied in situ with the use of two markers: the ultrastructural presence of Birbeck-like granules and immunohistochemistry with an antibody against human langerin/CD207 that cross-reacts with salmonid tissues. The appearance of Birbeck-like granules was observed in rainbow trout at 2 weeks post-hatch (PH) in the thymus and anterior kidney prior to the development of the spleen. Spleen first appeared at 3 weeks PH in both Atlantic salmon and rainbow trout, and Birbeck-like granules were observed within cells of the newly developed spleens. The cross-reactivity of langerin as seen by immunohistochemistry was not clearly observed in kidney and spleen until 9 weeks PH, when a strong cytoplasmic reaction was observed. To study langerin-positive cells in spleen and kidney during disease, microsporidial gill disease (MGD) in rainbow trout was used as a known disease model inducing a strong cell-mediated adaptive immune response. Langerin-positive cells in healthy fish were seen predominantly in the spleen, and only low numbers were present in the anterior kidney. During MGD, langerin-positive cell numbers were elevated in the anterior kidney and were significantly higher during 5, 6, and 10 weeks post-exposure (PE) compared with healthy control tissue. During MGD, the distribution of langerin-positive cells in the spleen and anterior kidney shifted from having significantly higher numbers of cells in the spleen than in the kidney in controls and at 1 and 4 weeks PE to having a similar distribution of the cells in the two organs at 2, 3, 5, and 6 weeks PE. By 10 weeks PE, significantly higher numbers of langerin-positive cells occurred in the anterior kidney compared with the spleen.

A case report of a localized reticular erythematous mucinosis like eruption of the lower legs mimicking cutaneous larvae migrans.

Reticular erythematous mucinosis is an uncommon type of cutaneous mucinosis. Typically, reticular erythematous mucinosis affects middle-aged women and usually appears as papules and plaques on the mid-upper trunk. Histologically, biopsies of reticular erythematous mucinosis reveal increased deposition of dermal mucin and perivascular lymphocytic inflammation. Infrequently, reticular erythematous mucinosis has been reported in males and in atypical locations other than the trunk. In these instances, reticular erythematous mucinosis can present a diagnostic challenge clinically and histologically. This case describes the clinical and histologic findings of a localized variant of reticular erythematous mucinosis in a 65-year-old male patient.

Primary Hyperoxaluria Type 1 (PH1) Presenting With End-Stage Kidney Disease and Cutaneous Manifestations in Adulthood: A Case Report.

RATIONALE: Primary hyperoxaluria (PH) is a rare autosomal recessive disorder more commonly diagnosed in children or adolescents. Owing to its rarity and heterogeneous phenotype, it is often underrecognized, resulting in delayed diagnosis, including diagnosis after end-stage kidney disease (ESKD) has occurred or recurrence after kidney-only transplantation. CASE PRESENTATION: A 40-year-old Caucasian Canadian woman with a history of recurrent nephrolithiasis since age 19 presented with ESKD and cutaneous symptoms. She had no known prior kidney disease and no family history of kidney disease or nephrolithiasis. DIAGNOSIS: A diagnosis of primary hyperoxaluria type 1 (PH1) due to homozygous splice donor mutation (AGXT c.680+1G>A) was made with kidney and cutaneous pathology demonstrating calcium oxalate deposition and ultrasound suggestive of nephrocalcinosis. INTERVENTIONS: She was initiated on frequent, high-efficiency, high-flux conventional hemodialysis and oral pyridoxine. Lumasiran was added 11 months later, after she developed bilateral swan-neck deformities. OUTCOMES: After 14 months of high-intensity dialysis and 3 months of lumasiran, there have been no signs of renal recovery, and extra-renal involvement has increased with progressive swan-neck deformities, reduced cardiac systolic function, and pulmonary hypertension. The patient has been waitlisted for kidney-liver transplantation. TEACHING POINTS: This case report describes an adult presentation of PH1. The case highlights the importance of timely workup of metabolic causes of recurrent nephrolithiasis or nephrocalcinosis in adults which can be a presenting sign of PH and genetic testing for PH to facilitate early diagnosis and treatment especially in the era of novel therapeutics that may alter disease course and outcomes. The case also demonstrates the value of testing for PH in adults presenting with unexplained ESKD and a history of recurrent nephrolithiasis or nephrocalcinosis due to implications for organ transplantation strategy and presymptomatic family screening.

JUSTIFICATION: L'hyperoxalurie primaire (HP) est un trouble récessif autosomique rare plus souvent rencontré chez les enfants ou les adolescents. En raison de sa rareté et de son phénotype hétérogène, cette affection est fréquemment sous-reconnue, ce qui entraîne un retard dans le diagnostic, et ce, même après l'apparition d'une insuffisance rénale terminale (IRT) ou une récidive suivant une greffe simple de rein. PRÉSENTATION DU CAS: Nous présentons le cas d'une Canadienne de race blanche âgée de 40 ans avec des antécédents de néphrolithiase récurrente depuis l'âge de 19 ans. La patiente était atteinte d'IRT et présentait des symptômes cutanés. Elle n'avait aucun antécédent connu de maladie rénale ou antécédent familial de maladie rénale ou de néphrolithiase. DIAGNOSTIC: Une pathologie rénale et cutanée montrant des dépôts d'oxalate de calcium et une échographie suggérant une néphrocalcinose ont permis de poser un diagnostic d'hyperoxalurie primaire de type 1 (HP1) due à une mutation de donneur d'épissage homozygote (AGXT c.680+1G>A). INTERVENTIONS: La patiente a amorcé des traitements d'hémodialyse conventionnelle à grande fréquence, à haut rendement et à flux élevé, et a reçu de la pyridoxine par voie orale. Un traitement par lumasiran a été ajouté 11 mois plus tard, après le développement de déformations bilatérales en col de cygne. RÉSULTATS: Après quatorze mois de dialyze à haute intensité et trois mois de lumasiran, aucun signe de récupération rénale n'a été observé. L'intervention d'épuration extra-rénale a été augmentée en raison de déformations progressives en col de cygne, d'une réduction de la fonction cardiaque systolique et d'une hypertension pulmonaire. La patiente a été placée sur la liste d'attente pour une transplantation rénale et hépatique. ENSEIGNEMENTS TIRÉS: Ce rapport de cas décrit une présentation adulte d'HP1. Ce cas souligne l'importance de traiter rapidement les causes métaboliques de la néphrolithiase ou de la néphrocalcinose récidivante chez les adultes, car celles-ci peuvent être des signes d'hyperoxalurie primaire (HP). Ce cas souligne en outre l'importance de procéder à des tests génétiques pour l'HP afin de permettre le diagnostic et le traitement précoces, en particulier à l'ère de nouveaux traitements susceptibles d'infléchir l'évolution et les résultats de la maladie. Enfin, il démontre la valeur du dépistage de l'HP chez les adultes présentant une IRT inexpliquée et des antécédents de néphrolithiase ou de néphrocalcinose récidivante, en raison de ses implications sur la stratégie de transplantation d'organes et sur le dépistage pré-symptomatique de la famille.

A case of multifocal cutaneous alternariosis in a patient with hairy cell leukemia.

We present a case of multifocal cutaneous alternariosis in a 69-year-old man with hairy cell leukemia. The patient presented with painful, violaceous nodules of the lower extremities and left forearm. His physical examination was otherwise normal. Biopsy specimens were submitted first for histopathological examination, followed by microbiological testing. Histopathology showed epidermal hyperplasia, intraepidermal abscess formation, and intracellular and extracellular fungal elements. Subsequent microbiological investigations, including fungal culture and polymerase chain reaction (PCR), isolated Alternaria alternata. The patient was treated for cutaneous alternariosis with itraconazole for 12 weeks and demonstrated significant improvement of the lesions. This case highlights the role of dual histopathological and microbiological laboratory investigation in suspected opportunistic cutaneous mycoses.

Les chercheurs présentent un cas d'alternariose cutanée multifocale chez un homme de 69 ans atteint d'une leucémie à tricholeucocytes. Le patient a consulté à cause de nodules violacés et douloureux sur les extrémités inférieures et l'avant-bras gauche. Son examen physique était autrement normal. Des biopsies ont été d'abord soumises en vue d'un examen histopathologique, puis de tests microbiologiques. L'histopathologie a révélé une hyperplasie épidermique, la formation d'abcès épidermiques et des éléments fongiques intracellulaires et extracellulaires. Les explorations microbiologiques subséquentes, y compris les cultures fongiques et l'amplification en chaîne par polymérase, ont isolé l'Alternaria alternata. Le patient a reçu un traitement de 12 semaines à l'itraconazole pour vaincre une alternariose cutanée, et ses lésions ont considérablement diminué. Ce cas souligne l'intérêt de conjuguer l'exploration histopathologique à l'exploration microbiologique dans les cas présumés de mycose opportuniste.

Infrared spectroscopy of synovial fluid as a potential screening approach for the diagnosis of naturally occurring canine osteoarthritis associated with cranial cruciate ligament rupture.

Objective: To evaluate infrared (IR) spectroscopy of synovial fluid (SF) as tool to differentiate between knees of dogs with naturally occurring OA associated with cranial cruciate ligament rupture (CrCLR) and controls. Method: 104 adult dogs with CrCLR (affected group) and 50 adult control dogs were recruited in a prospective observational study. Synovial fluid (SF) samples were collected preoperatively from dogs with CrCLR and from a subset of these at 4-, and 12-week post-surgery. Knee samples were collected bilaterally once from control dogs. Dried synovial fluid films were made, and IR absorbance spectra acquired. After preprocessing, partial least squares discriminant analysis (PLS-DA) and ANOVA-simultaneous component analysis (ASCA) were used to evaluate group and temporal differences, and to develop predictive models. Results: There were statistically significant spectral differences between the SF of OA affected and control dogs at all three time-points (P < 0.001). Pairwise comparison of spectral SF of knees with CrCLR over time showed statistically significant differences amongst all three time-points (P < 0.001). The predictive model for identifying the affected group from control had sensitivity, specificity and overall accuracy of 97.6%, 99.7% and 98.6%, respectively. Conclusions: The findings demonstrate the ability of FTIR-spectroscopy of synovial fluid combined with chemometric methods to accurately differentiate dogs with OA secondary to CrCLR from controls. The role of this IR-based screening test as a diagnostic and monitoring biomarker for OA specific to the joint being sampled warrants further investigation.

Methicillin-resistant Staphylococcus aureus replication in the presence of high (≥32 µg/ml) drug concentration of vancomycin as seen by electron microscopy.

Methicillin-resistant Staphylococcus aureus (MRSA) has unfortunately become a common pathogen in many healthcare facilities. In many institutions, vancomycin remains the preferred agent for treating serious MRSA infections including bacteraemia with or without endocarditis. The mutant prevention concentration (MPC) testing ≥109 colony forming units of bacteria, describes the antimicrobial drug concentration blocking the growth of the least susceptible cell from high density bacterial populations. With blood culture isolates of MRSA, we discovered strains with MPC values ≥32 µg/ml and viable cells could be readily recovered from agar plates containing 32 µg/ml of vancomycin. To investigate MRSA strains surviving in high concentrations of vancomycin on drug containing agar plates, we utilized electron microscopy to measure cell wall thickness as this has been previously reported as a potential mechanism of resistance1 along with septum thickening. Our data shows MRSA replication from high density bacterial populations in the presence of ≥32 µg/ml of vancomycin. Such observations may explain vancomycin failure in some patients and/or persistent bacteraemia and could potentially question the use of this drug in some critically ill patients in favour of an alternative agent.

Langerin/CD207 positive dendritic-like cells in the haemopoietic tissues of salmonids.

The presence of dendritic cells in fish is studied with immunohistochemistry using a commercially available antibody developed against Langerin/CD207 present in human Langerhans cells. Langerin/CD207, a protein known to be associated with the development of Birbeck granules in human and murine systems, was found to be expressed within the cytoplasm of spleen and head kidney cells of Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss). Reactivity was also observed within a few number of cells within the head kidney of Atlantic salmon, but not observed in any other tissues examined. Immunohistochemical results showed Langerin/CD207 reactivity in the cytoplasm of cells in Atlantic salmon and rainbow trout comparable to reactivity seen in human Langerhans cells. The results in this study further corroborate the presence of dendritic cells with remarkable similarities to human Langerhans cells in the spleens and to a lesser extent in head kidney of salmonids.

Infectious salmon anaemia virus replication and induction of alpha interferon in Atlantic salmon erythrocytes.

BACKGROUND: Infectious salmon anaemia (ISA) virus (ISAV), which causes ISA in marine-farmed Atlantic salmon, is an orthomyxovirus belonging to the genus Isavirus, family Orthomyxoviridae. ISAV agglutinates erythrocytes of several fish species and it is generally accepted that the ISAV receptor destroying enzyme dissolves this haemagglutination except for Atlantic salmon erythrocytes. Recent work indicates that ISAV isolates that are able to elute from Atlantic salmon erythrocytes cause low mortality in challenge experiments using Atlantic salmon. Previous work on ISAV-induced haemagglutination using the highly pathogenic ISAV strain NBISA01 and the low pathogenic ISAV strain RPC/NB-04-0851, showed endocytosis of NBISA01 but not RPC/NB-04-0851. Real-time RT-PCR was used to assess the viral RNA levels in the ISAV-induced haemagglutination reaction samples, and we observed a slight increase in viral RNA transcripts by 36 hours in the haemagglutination reaction with NBISA01 virus when the experiment was terminated. However, a longer sampling interval was considered necessary to confirm ISAV replication in fish erythrocytes and to determine if the infected cells mounted any innate immune response. This study examined the possible ISAV replication and Type I interferon (IFN) system gene induction in Atlantic salmon erythrocytes following ISAV haemagglutination. RESULTS: Haemagglutination assays were performed using Atlantic salmon erythrocytes and one haemagglutination unit of the two ISAV strains, NBISA01 and RPC/NB-04-0851, of differing genotypes and pathogenicities. Haemagglutination induced by the highly pathogenic NBISA01 but not the low pathogenic RPC/NB-04-0851 resulted in productive infection as evidenced by increased ISAV segment 8 transcripts and increase in the median tissue culture infectious dose (TCID50) by 5 days of incubation. Moreover, reverse transcription (RT) quantitative PCR used to compare mRNA levels of key Type I IFN system genes in erythrocyte lysates of haemagglutination reactions with the two ISAV strains showed a higher relative fold increase of IFN-alpha in NBISA01 haemagglutinations compared to RPC/NB-04-085-1 haemagglutinations (33.0 - 44.26 relative fold increase compared to 11.29). Erythrocytes exposed to heat-inactivated virus or to polyinosinic:polycytidylic acid (polyI:C) or to L-15 medium alone (negative control assays) had minimal late induction (<3.5 relative fold increase) of STAT1 and/or ISG15 and Mx genes, whereas erythrocytes exposed to UV-inactivated virus lacked any cytokine induction. CONCLUSION: ISAV-induced haemagglutination by a highly pathogenic virus strain results in virus uptake and productive infection of Atlantic salmon erythrocytes accompanied by significant induction of IFN-alpha. This study also highlights the critical role of ISAV strain variation in the initial stages of the virus-cell interaction during haemagglutination, and possibly in the pathogenesis of ISA. Moreover, the study shows for the first time that fish erythrocytes immunologically respond to ISAV infection.

Demonstration of infectious salmon anaemia virus (ISAV) endocytosis in erythrocytes of Atlantic salmon.

Infectious salmon anaemia (ISA) virus (ISAV) is a fish orthomyxovirus that has recently been assigned to the new genus Isavirus within the family Orthomyxoviridae. It possesses the major functional characteristics of the virus family including haemagglutinating, receptor destroying enzyme (RDE), and fusion activities associated with the virion surface proteins. It is generally accepted that ISAV agglutinates erythrocytes of several fish species and that the ISAV RDE activity dissolves this haemagglutination reaction except for Atlantic salmon (Salmo salar) erythrocytes. We used electron microscopy to examine the physical interaction between ISAV and erythrocytes from Atlantic salmon and rainbow trout (Oncorhynchus mykiss) during haemagglutination. We present evidence that ISAV enters into Atlantic salmon erythrocytes. Atlantic salmon erythrocytes incubated with ISAV for 4 hours showed endocytosis of the virus particles, which is consistent with virus infection. These observations suggest that the lack of dissolution of ISAV-induced haemagglutination of Atlantic salmon erythrocytes favours virus infection of the erythrocytes. Moreover, such a haemagglutination-infection phenotype is fundamentally different from haemagglutination by avian and mammalian orthomyxoviruses, and is indicative of a different pathogenesis for the fish orthomyxovirus.

In vitro heterogeneity of osteogenic cell populations at various equine skeletal sites.

Bone cell cultures were evaluated to determine if osteogenic cell populations at different skeletal sites in the horse are heterogeneous. Osteogenic cells were isolated from cortical and cancellous bone in vitro by an explant culture method. Subcultured cells were induced to differentiate into bone-forming osteoblasts. The osteoblast phenotype was confirmed by immunohistochemical testing for osteocalcin and substantiated by positive staining of cells for alkaline phosphatase and the matrix materials collagen and glycosaminoglycans. Bone nodules were stained by the von Kossa method and counted. The numbers of nodules produced from osteogenic cells harvested from different skeletal sites were compared with the use of a mixed linear model. On average, cortical bone sites yielded significantly greater numbers of nodules than did cancellous bone sites. Between cortical bone sites, there was no significant difference in nodule numbers. Among cancellous sites, the radial cancellous bone yielded significantly more nodules than did the tibial cancellous bone. Among appendicular skeletal sites, tibial metaphyseal bone yielded significantly fewer nodules than did all other long bone sites. This study detected evidence of heterogeneity of equine osteogenic cell populations at various skeletal sites. Further characterization of the dissimilarities is warranted to determine the potential role heterogeneity plays in differential rates of fracture healing between skeletal sites.

Developmental transformations in a normal series of embryos of the sea lamprey Petromyzon marinus (Linnaeus).

Lamprey development is of interest to evolutionary biologists because it can inform our understanding of primitive vertebrate developmental patterns. In this study, we describe and illustrate some of the principle landmarks of organogenesis in the embryonic sea lamprey Petromyzon marinus L. at different chronological ages. We examined 63 fixed embryos spanning Piavis developmental stages 11-18+ (5-70 days postfertilization) by gross observation and histology. This period begins at late neurulation stages and ends with the formation of the larva (ammocoete). A significant difference with some previous accounts is that the anus develops not from a persistent blastopore, but by secondary canalization and proctodeum formation at the former site of the blastopore. Further, we show that the ciliated bands of the pharyngeal roof originate in the esophagus, distinguishing it from the intestine. We clarify the epithelialization of the gut, showing that the secondary gut cavity is progressively epithelialized from each end. We identify possible germ cells in the coelomic and cloacal walls. Balfour's "subnotochordal rod" is lacking in our specimens; we suggest that he may have misinterpreted the corpus adiposum. Our study is of potential value to the growing number of biologists interested in lamprey development and provides a character set that will be used : 1) in a phylogenetic study of vertebrate development, and 2) to prepare a staging series for the lamprey based on parsimony analysis.

Ultrastructural evidence of autoinfection in the gills of Atlantic cod Gadus morhua infected with Loma sp. (phylum Microsporidia).

Infection by a microsporidian of the genus Loma was found in gills of cod Gadus morhua. Xenomas contained parasites in multiple stages of development. Some spores looked empty and had everted polar tubes, which were either straight or coiled. These polar tubes were scattered throughout the xenoma cytoplasm, and some of them pierced the plasma membrane. Those outside of the xenoma penetrated neighboring cells, including blood cells. These observations suggest that a mechanism of autoinfection could occur in blood cells and gill tissue, perpetuating the disease in the host.