Cardiotoxicity, inflammation, and immune response after rattlesnake envenomation in the horse.

BACKGROUND: Cardiac abnormalities are reported in rattlesnake-bitten horses. The prevalence and cause are unknown. OBJECTIVES: To detect cardiac damage in rattlesnake-bitten horses by measuring cardiac troponin I (cTnI) and evaluating ECG recordings for presence of arrhythmias, and explore causes of this cardiac damage by measuring venom excretion, anti-venom antibodies, and tumor necrosis factor alpha (TNFα). ANIMALS: A total of 20 adult horses with a clinical diagnosis of rattlesnake bite and 6 healthy adult horses. METHODS: In a prospective clinical study, bite site swabs, blood samples, and urine samples were collected at various time points from 20 horses with a clinical diagnosis of snake bite. Continuous ECG recordings were obtained on the 20 affected horses and 6 normal control horses using 24-hour holter monitors. Plasma samples were assayed for cTnI, serum samples were assayed for TNFα and anti-venom antibodies, and bite site swabs and urine were assayed for venom. RESULTS: Forty percent of rattlesnake-bitten horses (8/20) experienced myocardial damage (increased cTnI). Seventy percent (14/20) experienced a cardiac arrhythmia. There was a positive correlation between cTnI and TNFα (P < .02). Horses with cTnI ≥ 2 ng/mL were more likely to have antibody titers >5,000 (P < .05). No correlations were found between venom concentration and cTnI, anti-venom antibody titers, TNFα, or presence of arrhythmias. CONCLUSIONS AND CLINICAL IMPORTANCE: Cardiac abnormalities in this population of horses indicate that cardiac damage after rattlesnake bite is common. Rattlesnake-bitten horses should be monitored for signs of cardiac damage and dysfunction. Long-term follow-up should be encouraged to detect delayed cardiac dysfunction.

Medulloblastoma/primitive neuroectodermal tumour studied as a Matrigel enhanced subcutaneous xenograft model.

An important role for pre-clinical models of medulloblastoma/primitive neuroectodermal tumour (MB/PNET) is inhibited by the limitations of conventional heterotransplantation. Nine cohorts of MB/PNET were studied for subcutaneous engraftment in nude mice by both conventional and Matrigel supplemented methods. While no subcutaneous tumours resulted from 63 conventional attempts, an aggregate 41 xenografts from 72 injections (57%) were produced when Matrigel was added to the cell suspension. In subsequent passage, engraftment rate approached 100%. To study the response to chemotherapeutic agents in the model, a total of 221 tumours in 3 cohorts were treated using one of the following: cisplatin, carboplatin, vincristine, cyclophosphamide, diaziquone, or saline control. While all agents demonstrated statistically significant activity, cyclophosphamide proved to be particularly effective. The potential applications of this xenograft model in the biologic as well as therapeutic study of MB/PNET deserve continuing investigation.

Failure of iron chelators to reduce tumor growth in human neuroblastoma xenografts.

Neuroblastoma (NB) is a high risk tumor of childhood, and raised serum ferritin is an adverse prognostic factor. The hypothesis that iron chelation therapy impacts tumor status and patient prognosis through changes in iron metabolism has been systematically evaluated here in a xenograft model of human NB. One of two iron chelators was given in seven different regimens to nude mice xenografted s.c. with either IMR-32, an established cell line, or JBN-1, heterotransplanted directly from a patient. Nude mice (a total of 160 in 24 cohorts) were given: desferrioxamine (DFO) by s.c. bolus or reservoir; 1,2-dimethyl-3-hydroxypyridin-4-one (L1), i.p. or orally; or saline. Measurements of mean Hb and liver iron levels were compared with corresponding saline cohorts per regimen as well as for pooled cohorts per agent for both cell lines. For IMR-32 xenografts, significant differences in Hb were achieved with L1 (10.9 g/dl pooled versus 13.7 g/dl controls) and in liver iron with DFO and L1 (235 microg/g and 306 microg/g, respectively, versus 520 microg/g). For JBN-1, the pattern was similar. With L1, H6 was 10.2 g/dl and controls were 11.7 g/dl (individual DFO cohorts were also significant); liver iron with DFO was 303 microg/g, liver iron with L1 was 270 microg/g, and controls were 387 microg/g. Additional therapy prior to tumor injection (67 mice and 10 cohorts) did not increase the depletion. Despite documentation of iron depletion, no reductions in tumor engraftment, latency, or tumor size at end point were achieved in the chelator-treated mice, compared with controls populations. Accordingly, inclusion of these iron chelators in clinical trials for NB appears unwarranted.

Response of brain tumors to chemotherapy, evaluated in a clinically relevant xenograft model.

Chemotherapy for brain tumors remains unsatisfactory. Despite increasing participation in clinical trials, there is a clear need for pre-clinical models. Heterotransplantation of surgical specimens directly into the anterior chamber of the nude mouse eye has been demonstrated to produce evaluable xenografts. Drug access in this model is considered to mimic the blood-brain barrier. Five clinical specimens in 3 children with primitive neuroectodermal tumor/medulloblastoma were the sources of 293 intraocular xenografts (5 cohorts by source). Each tumor-bearing mouse received 1 of 5 drugs or normal saline, by intraperitoneal injection, weekly for 5 weeks. Response was monitored for up to 22 weeks, using a staging system which estimates the proportion of the anterior chamber filled by tumor. Results were analysed both as response rates (shrinkage in excess of 50%) at the conclusion of the treatment course and as time to tumor progression by the life table method. Comparison of response rates within cohorts by source of xenografts (exact chi-square test for overall and 2-sided Fisher's exact test for paired comparisons) indicated cyclophosphamide to be the most effective single agent. In logrank analyses cyclophosphamide achieved significantly longer delays to progression than all other drugs in one cohort and longer delays than all but diaziquone in 2 other cohorts. The intraocular xenograft model is a clinically relevant system for the study of therapeutic agents in brain tumors. The effectiveness of intensive dosage cyclophosphamide in a model dependent on access across the blood-aqueous barrier is important and consistent with recent clinical data.

Heterotransplantation of early B-lineage acute lymphoblastic leukemia using a solubilized attachment matrix (Matrigel).

Maintenance of long term culture and conventional xenografting of early B-lineage acute lymphoblastic leukemia cells is most difficult. Matrigel, a solubilized attachment matrix shown to aid growth of anchorage dependent solid tumors, was studied in heterotransplantation. Material for xenografting was derived from 5 patient bone marrow aspirates and 5 cell lines previously established and maintained by intraocular inoculation in nude mice. Specimens were injected by 3 methods: intraocular (n = 397); s.c. in medium (n = 78); and s.c. in medium supplemented by Matrigel (n = 69). With intraocular injection, 6 of 10 cell sources grew with respective ingraftment rates of 29-76%. Using the conventional s.c. method, no tumors resulted. The addition of Matrigel produced s.c. ingraftment from 8 of 10 cell sources (ingraftment rate, 50-100%). Immunophenotype, histopathology, and karyotype of the cells derived after Matrigel dependent ingraftment correlated with the cells of origin. It is concluded that Matrigel enables establishment and maintenance of early B-lineage acute lymphoblastic leukemia cell growth in a s.c. xenograft model.