Biomedical subjects
Publications and source records attributed to J D Gilbert.
A case of unexpected death in a 65-year-old woman, caused by a migrating foreign body that resulted in multifocal hepatic abscesses, is reported. The foreign body was subsequently identified as a portion of chicken fibula. The prolonged time course of the illness, with relatively nonspecific symptoms and signs, resulted in establishment of the diagnosis only at autopsy.
Three cases of traditional punishment in Central Australian Aboriginal men are presented in which the thighs were speared or stabbed as part of a "payback" system. In two cases, an unexpected effect of the stabbing or spearing was death due to severing of major leg vessels. The relationship between customary Aboriginal law and general law in Australia has not been clearly defined; however, these cases demonstrate that significant and untoward effects may result from traditional punishments. Pathologists working near traditional Australian Aboriginal communities may still encounter such injuries at autopsy.
A retrospective study of cases of drowning suicide was undertaken at the Forensic Science Centre in Adelaide, South Australia for the period April 1980 to March 2000. A total of 123 cases were found, with 76 males (age, 16-88 years; average, 50.5 years; standard deviation [SD], 20.1 years) and 47 females (age, 34-88 years; average, 60.6 years; SD, 13.9 years). There were 66 fresh water drownings and 57 saltwater drownings. Female victims were significantly older than male victims for both fresh water and saltwater drownings (P < .05 and P < .01, respectively). Deaths in young women were rare. No temporal trend in drowning suicides could be shown, with annual numbers varying from 0 to 12 cases (average, 6.15; median, 7). Women preferentially chose the ocean or bath to drown themselves in, whereas males chose rivers, ditches, and lakes. Swimming pools were rarely used for suicide in this population; alcohol use was not usual; and there was often a significant history of mental illness.
A 32-year-old man who died suddenly and unexpectedly was found at autopsy to have prominent fatty infiltration of his left ventricle with fibrous scarring, in the presence of normal coronary arteries. The right ventricle was minimally involved. A diagnosis of ventricular dysplasia largely limited to the left ventricle was made. Subsequent family screening identified a brother with clinical manifestations of ventricular dysplasia. This case provides further evidence for the association of left ventricular dysplasia with sudden death, and demonstrates that left ventricular involvement may also be inheritable. Whether predominantly left ventricular dysplasia is a manifestation of right ventricular dysplasia, or is a separate entity, is yet to be determined.
A 20-year-old woman who suffered from septo-optic dysplasia died unexpectedly following a presumed viral gastroenteritis. Autopsy and neuropathological examination confirmed optic nerve hypoplasia with absence of the septum pellucidum. Marked adrenal gland hypoplasia reflected hypothalamic hypopituitarism. The small and large intestines were dilated and filled with liquid fecal material. This case demonstrates that individuals with septo-optic dysplasia may be at risk of unexpected death at all ages. The complexity of mechanisms of death in rare dysmorphic conditions may be overlooked if relevant clinical information is not available at the time of autopsy and unless specific steps are taken to clearly delineate the underlying features of the condition.
A rare complication of the use of glycine irrigation fluid during prostatic surgery in a 69-year-old man is described. Following cystolithopexy and transurethral resection of the prostate for benign prostatomegaly, abdominal distension developed with increasing ventilatory pressures. Despite retroperitoneal fluid evacuation at subsequent urgent laparotomy, cardiac arrest occurred that was not amenable to resuscitation. At autopsy a traumatic defect in the posterior bladder wall filled with calculus debris was confirmed that did not communicate with the peritoneal cavity. Hyponatremia with markedly elevated levels of blood, urine, and body fluid glycine were demonstrated. Death was, therefore, attributed to glycine toxicity following tracking of glycine through a surgical defect in the posterior bladder wall. Careful dissection of surgical sites is required in such cases to demonstrate any additional trauma that may be associated with the fatal episode. Analysis of body fluids for glycine and electrolytes is also necessary to assist in the determination of possible mechanisms of death.
The E1B-deleted, replication-competent ONYX-015 (dl1520) adenovirus was originally described as being able to selectively kill p53-deficient cells due to a requirement of p53 inactivation for efficient viral replication. This hypothesis has become controversial because subsequent in vitro studies have demonstrated that the host range specificity of ONYX-015 is independent of p53 gene status. Using a pair of isogenic cell lines that differ only in their p53 status, we demonstrate here that although ONYX-015 can replicate in both p53 wild-type and mutant cells in vitro, the virus demonstrates significantly greater antitumor activity against mutant p53 tumors in vivo. Moreover, ONYX-015 viral therapy can be combined with radiation to improve tumor control beyond that of either monotherapy. The results demonstrate that ONYX-015 can discern in vivo between tumors having a different p53 status and that it may be an effective neoadjuvant to radiation therapy.
Replication-competent adenoviruses may provide a highly efficient means of delivering therapeutic genes to tumors. Previously, we evaluated in vitro a replication-competent adenovirus (Ad5-CD/TKrep) containing a cytosine deaminase (CD)/herpes simplex type 1 thymidine kinase (HSV-1 TK) fusion gene that allows lytic viral therapy to be combined with double suicide gene therapy. Both the CD/5-FC and HSV-1 TK/GCV enzyme/prodrug systems enhanced the tumor cell-specific cytopathic effects of the Ad5-CD/TKrep virus in vitro and sensitized cells to radiation. To extend these in vitro findings in vivo, we evaluated the antitumor activity of the Ad5-CD/TKrep virus in combination with double prodrug therapy and radiation therapy. The Ad5-CD/TKrep virus independently demonstrated significant antitumor activity against C33A cervical carcinoma xenografts. Therapeutic outcome was dramatically improved with systemic administration of double, but not single, prodrug (5-FC + GCV) therapy. When used in a neoadjuvant setting, Ad5-CD/TKrep-mediated double suicide gene therapy dramatically potentiated the effectiveness of radiation therapy. The trimodal approach of Ad5-CD/TKrep viral, double suicide gene, and radiotherapies produced significant tumor regression and ultimately 100% tumor cure. The results demonstrate the high therapeutic potential of the trimodal approach and provide a solid foundation for future clinical trials.
Adenovirus-mediated gene transfer may hold much promise in the treatment of human cancer. However, concerns regarding vector dissemination beyond the target tissue, particularly with replication-competent viruses, require an evaluation of the persistence of viral infection in collateral tissue and vector-associated toxicities. In addition, for indications such as prostate cancer, the proximity of the point of viral administration to organs of the male reproductive system raises concerns regarding inadvertent germ-line transmission of genes carried by the virus. To address these concerns, the biodistribution, persistence, toxicity, and potential of germ-line transmission of a replication-competent adenovirus (Ad5-CD/TKrep) following intraprostatic administration in the mouse was examined. Ad5-CD/TKrep (10(10) vp, 5 x 10(11) vp/kg) was injected intraprostatically on Day 1 of the study and its presence in the major organs of the male urogenital tract (prostate, testes, seminal vesicles, and urinary bladder) and liver was determined on Days 8 and 29. For comparison, a parallel group of animals was injected with the same dose of a related replication-defective Ad5-FGNR virus. To evaluate germ-line transmission, Ad5-CD/TKrep-injected males were mated to females on Days 8 and 29 and resulting embryos were examined for AdS-CD/TKrep viral DNA. Ad5-CD/TKrep viral DNA was detected in all major organs of the adult male urogenital tract and liver 7 and 28 Days postinjection. Interestingly, relative to the replication-defective Ad5-FGNR adenovirus, the replication-competent Ad5-CD/TKrep virus accumulated to a much greater level (approximately 300-fold) and persisted for a longer period of time in prostate, testes, and liver. This difference could not be explained on the basis of differences in viral infectivity, suggesting that the AdS-CD/TKrep virus may be capable of replicating in mouse tissues in vivo. In vitro infection of six mouse cell lines representing prostate, testes, and liver demonstrated that the Ad5-CD/TKrep virus was indeed capable of replicating in these mouse cell types, albeit with reduced efficiencies relative to human cells. Despite the fact that the Ad5-CD/TKrep vector persisted in the adult male gonads and may have replicated in vivo, we observed no evidence of germ-line transmission in 149 offspring examined. To evaluate the toxicity of combining Ad5-CD/TKrep viral therapy with CD/5-FC and HSV-1 TK/GCV suicide gene therapies as a prerequisite for a human trial, an escalating dose (10(8), 10(9), 10(10) vp) of Ad5-CD/TKrep was administered intraprostatically followed by 7 days of 5-FC and GCV double prodrug therapy. Although the virus persisted in the mouse urogenital tract and liver for up to 28 days postinjection, most of the toxicities observed were expected, minimal, and self-limiting. These results lead us to believe that intraprostatic administration of the Ad5-CD/TKrep virus to humans concomitant with double suicide gene therapy will be associated with acceptable toxicities and will not result in vertical transmission of viral-encoded genes through the germ line.
A retrospective analysis of 68 cases of fatal childhood pedestrian accidents in South Australia that occurred over a 20-year period from January 1977 to December 1996 was undertaken. The age range was 12 months to 16 years (average, 7.6 years), with a male to female ratio of 41:27. The most common site of significant injury was the head (91.2%), followed by the abdomen (50%), chest (47.1%), and neck (38.2%). The most frequently encountered significant injury combinations involved the head, chest, and abdomen (14.7%); the head and chest (11.8%); and the head, neck, and abdomen (11.8%). Injuries were severe, consisting of extensive compound and comminuted skull fractures, neck fractures, and massive intraparenchymal brain and internal organ damage with avulsions and fragmentation. Non-life-threatening limb injuries occurred in 88% of cases. A distinct subgroup involved infants and younger children playing in driveways at home who were hit by reversing vehicles. The injuries were often of such a severe nature that death was instantaneous and with no possibility of successful medical intervention. More deaths occurred during the winter than summer months.
To examine the pattern of injuries in cases of fatal shark attack in South Australian waters, the authors examined the files of their institution for all cases of shark attack in which full autopsies had been performed over the past 25 years, from 1974 to 1998. Of the seven deaths attributed to shark attack during this period, full autopsies were performed in only two cases. In the remaining five cases, bodies either had not been found or were incomplete. Case 1 was a 27-year-old male surfer who had been attacked by a shark. At autopsy, the main areas of injury involved the right thigh, which displayed characteristic teeth marks, extensive soft tissue damage, and incision of the femoral artery. There were also incised wounds of the right wrist. Bony injury was minimal, and no shark teeth were recovered. Case 2 was a 26-year-old male diver who had been attacked by a shark. At autopsy, the main areas of injury involved the left thigh and lower leg, which displayed characteristic teeth marks, extensive soft tissue damage, and incised wounds of the femoral artery and vein. There was also soft tissue trauma to the left wrist, with transection of the radial artery and vein. Bony injury was minimal, and no shark teeth were recovered. In both cases, death resulted from exsanguination following a similar pattern of soft tissue and vascular damage to a leg and arm. This type of injury is in keeping with predator attack from underneath or behind, with the most severe injuries involving one leg. Less severe injuries to the arms may have occurred during the ensuing struggle. Reconstruction of the damaged limb in case 2 by sewing together skin, soft tissue, and muscle bundles not only revealed that no soft tissue was missing but also gave a clearer picture of the pattern of teeth marks, direction of the attack, and species of predator.
Retrospective review was undertaken of all autopsies in which methadone was detected at the Forensic Science Centre, South Australia, during a 3-year period from July 1996 to June 1999. Thirty-five cases were found in which methadone had either caused or contributed to death (age range = 14-54 years; average = 31 years; M:F = 3.4:1). Ten victims were participating in a methadone maintenance program, of whom four died within a week of enrollment. Eight victims (23%) not enrolled in a methadone maintenance program were found who had died after the use of "diverted" methadone (i.e., prescribed for someone else) (age range = 14-34 years; average = 25 years; M:F = 6:2). Deaths were directly attributable to methadone toxicity in seven of the eight cases, with additional drugs or alcohol being found in seven cases. Prevention of ongoing deaths caused by methadone diversion could be achieved by allowing only daily releases of methadone, with the addict having to consume the drug under close supervision.
Several nonsedating histamine H1-receptor antagonists are associated with torsades de pointes ventricular tachycardia. The objectives of this study were to: (i) compare electrocardiographic, monophasic action potential, and arrhythmogenic effects of sedating and nonsedating H1-receptor antagonists, and (ii) identify correlates of drug-induced torsades de pointes in an isolated ventricle model. Isolated, electrically paced (1-3 Hz) rabbit ventricles were Langendorff-perfused with either drug-free Tyrode's solution or one of the following: (i) the sedating H1-receptor antagonist hydroxyzine (0.1-30 microM), (ii) cetirizine, a nonsedating metabolite of hydroxyzine (1-300 microM), and (iii) the nonsedating, putatively arrhythmogenic H1-receptor antagonist astemizole (0.1-30 microM). Volume conducted electrocardiographic signals and monophasic action potentials from the periapical left ventricular endocardium and epicardium were recorded. There were no apparent changes in control (n = 15) or hydroxyzine-perfused (n = 7) hearts. Cetirizine (n = 13) produced a mild biphasic electrocardiographic QT interval prolongation and was associated with early afterdepolarizations, but not with torsades de pointes. Astemizole (n = 11) lengthened QT intervals, and at high concentration (30 microM) induced torsades de pointes in 10 of 11 hearts (P < 0.001 vs. all other groups). These findings are consistent with previously reported repolarizing current inhibition by cetirizine, but may additionally indicate "compensatory" inhibition of inward currents at higher concentrations. By contrast, astemizole-induced changes are consistent with unopposed repolarizing current inhibition.
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Recent evidence has suggested that tumor cells having a wild-type p53 status are more sensitive to chemotherapeutic agents and radiation than cells that lack functional p53. The heightened sensitivity of wild-type p53 cells is thought to be attributable to their propensity to undergo p53-mediated apoptosis after insult. Given that suicide gene therapy is essentially tumor-targeted chemotherapy, we examined the hypothesis that coexpression of wild-type p53 could enhance the efficacy of adenovirus-mediated suicide gene therapy. Human Hep3B and SK-OV-3 cells, which are null for p53, were infected with a pair of replication-deficient adenoviruses that expressed a cytosine deaminase/herpes simplex virus thymidine kinase (CD/HSV-1 TK) fusion gene without (fusion gene nonreplicative adenovirus, FGNR) or with (FGNRp53) the wild-type human p53 gene. The sensitivity of cells to the CD/5-fluorocytosine (CD/5-FC) and HSV-1 TK/ ganciclovir (GCV) enzyme/prodrug systems was determined in vitro and in vivo. Coexpression of p53 did not enhance the cytotoxicity of either the CD/5-FC or HSV-1 TK/GCV system in vitro. The failure to observe an effect of p53 could not be explained on the basis of insufficient or transient p53 expression, because FGNRp53-infected cells growth arrested in G1, induced Bax, and underwent apoptosis at an increased rate after prodrug treatment, particularly when the adenovirus E1A protein was present. Intratumoral injection of FGNRp53 concomitant with single or double pro-drug therapy resulted in a tumor growth delay that was equal to or less than that observed with the FGNR virus. Our results indicate that coexpression of p53 may not necessarily improve the efficacy of adenovirus-mediated CD/ 5-FC and HSV-1 TK/GCV suicide gene therapies in vivo.
Human translocation liposarcoma (TLS)-CCAAT/enhancer binding protein (C/EBP) homologous protein (CHOP) is a fusion oncoprotein found specifically in a malignant tumor of adipose tissue and results from a t(12;16) translocation that fuses the amino-terminal part of TLS to the entire coding region of CHOP. Being that CHOP is a member of the C/EBP transcription factor family, proteins that comprise part of the adipocyte differentiation machinery, we examined whether TLS-CHOP blocked adipocyte differentiation by directly interfering with C/EBP function. Using a single-step retroviral infection protocol, either wild-type or mutant TLS-CHOP were co-expressed along with C/EBPbeta in naïve NIH3T3 cells, and their ability to inhibit C/EBPbeta-driven adipogenesis was determined. TLS-CHOP was extremely effective at blocking adipocyte differentiation when expressed at a level comparable to that observed in human myxoid liposarcoma. This effect of TLS-CHOP required a functional leucine zipper domain and correlated with its ability to heterodimerize with C/EBPbeta and inhibit C/EBPbeta DNA binding and transactivation activity in situ. In contrast, the TLS-CHOP basic region was dispensable, making it unlikely that the inhibitory effect of TLS-CHOP is attributable to unscheduled gene expression resulting from TLS-CHOP's putative transactivation activity. Another adipogenic transcription factor, PPARgamma2, was able to rescue TLS-CHOP-inhibited cells, indicating that TLS-CHOP interferes primarily with C/EBPbeta-driven adipogenesis and not with other requisite events of the adipocyte differentiation program. Together, the results demonstrate that TLS-CHOP blocks adipocyte differentiation by directly preventing C/EBPbeta from binding to and transactivating its target genes. Moreover, they provide strong support for the thesis that a blockade to normal differentiation is an important aspect of the cancer process.
Two obstacles limiting the efficacy of nearly all cancer gene therapy trials are low gene transduction efficiencies and the lack of tumor specificity. Recently, a replication-competent, E1B-attenuated adenovirus (ONYX-015) was developed that could overcome these limitations, because it was capable of efficiently and selectively destroying tumor cells lacking functional p53. In an attempt to improve both the efficacy and safety of this approach, we constructed a similar adenovirus (FGR) containing a cytosine deaminase (CD)/herpes simplex virus type-1 thymidine kinase (HSV-1 TK) fusion gene, thereby allowing for the utilization of double-suicide gene therapy, which has previously been demonstrated to produce significant antitumor effects and potentiate the therapeutic effects of radiation. The FGR virus exhibited the same tumor cell specificity and replication kinetics as the ONYX-015 virus in vitro. Importantly, both the CD/5-FC and HSV-1 TK/GCV suicide gene systems markedly enhanced the tumor cell-specific cytopathic effect of the virus, and, as expected, sensitized tumor cells to radiation. By contrast, neither the FGR virus nor either suicide gene system showed significant toxicity to normal human cells. Both suicide gene systems could be used to suppress viral replication effectively, thereby providing a means to control viral spread. The results support the thesis that the three-pronged approach of viral therapy, suicide gene therapy, and radiotherapy may represent a powerful and safe means of selectively destroying tumor cells in vivo.
Assays have been established for the quantitation of timolol and its 13C3- and 2H9-stable-isotope-labeled analogs in plasma and urine using liquid chromatography with atmospheric-pressure chemical-ionization tandem mass spectrometry. For the analysis of urine, underivatized timolol and its labeled analogs are monitored while timolol in plasma is assayed down to concentrations of 0.2 ng/mL after derivatization with phosgene. The great power of this technique is illustrated by simultaneously assaying three different species of timolol in plasma and urine obtained from dogs receiving simultaneous ophthalmic, oral, and intravenous doses of unlabeled and [2H9]- and [13C3]-labeled timolol. Thus, the ophthalmic and oral bioavailabilities of timolol are measured in a single experiment rather than as a three-phase crossover experiment. This approach yields accurate and precise analytical data, obviates intrasubject variability, and saves both analytical and animal resources.