Y2k revisited: a human component?
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Attitude to Death↗
Biomedical subjects
J H Sinard
Publications and source records attributed to J H Sinard.
Factors affecting autopsy rates, autopsy request rates, and autopsy findings at a large academic medical center.
Autopsy rates continue to decline in the United States. To assess the impact of various objective factors (time of death, day of death, age at death, patient gender, clinical service, and length of hospital stay) on the autopsy request rate, autopsy rate, successful request rate, and percentage of cases in which the autopsy examination added to or altered the clinical assessment of the patient, we prospectively studied all hospital deaths at a major academic medical center for the 3-year period from 1996 through 1998. The autopsy rate decreases significantly with patient age, both because of a decreasing request rate and because family members are less likely to grant permission. An autopsy is less likely to be requested for deaths in the emergency department or on general surgery services and most likely to be requested for fetal, medicine, cardiothoracic surgery, and pediatric deaths. Families more commonly grant permission for autopsy on fetal deaths, pediatric deaths, and emergency department deaths. Forty percent of autopsies reveal significant information about the patient's death beyond what was known premortem. This is least frequent among the fetal deaths, but relatively constant for adults of all ages. Patients who die in the emergency department are most likely to have significant unexpected findings at autopsy. Increasing the request rate for adult patients who die in the emergency department and on the medicine services will result in the greatest increase in information learned from autopsy.
Age Distribution↗
Informatics and anatomic pathology: meeting challenges and charting the future.
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Humans↗
Quality improvement on an academic autopsy service.
CONTEXT: Autopsy rates continue to decline in the United States. OBJECTIVE: Although many of the causes of this decline are external to pathology departments, we hypothesized that intradepartmental efforts to improve the quality of the service we provide to our clinical colleagues could increase our autopsy rate. METHOD: We developed a multifaceted quality improvement program for our autopsy service aimed at increasing the visibility of the service, improving the service's reporting, and increasing the amount and quality of data available from the service. SETTING: A large academic medical center that performs approximately 250 autopsies each year. RESULTS: After implementation of our quality improvement program, the decline in our autopsy rate has not only stopped, but rates have even begun to increase. Additionally, physician satisfaction surveys conducted before and after implementation of our quality improvement initiatives showed an across-the-board improvement in clinician perception of the service. CONCLUSION: Pathologists can and should be proactive in addressing the declining autopsy rate, rather than viewing it as someone else's problem or hoping that someone else will protect this important quality assurance tool for medical care.
Attitude of Health Personnel↗
Implementation of a practical digital imaging system for routine gross photography in an autopsy environment.
BACKGROUND: The autopsy environment places stringent requirements on a digital imaging system. These requirements must be addressed if the system is to be functional, easy to use, and reliable. DESIGN: After clearly defining the requirements for such a system, we implemented routine digital imaging in a busy academic autopsy suite. RESULTS: The new technology was immediately accepted by both the resident staff and the technical staff. Although a 35-mm camera was always available for traditional photography, it was rarely used. An interesting side effect of implementing digital imaging was a nearly twofold increase in the number of images taken per autopsy case. The requirements, features, and utility of a digital imaging system are discussed. CONCLUSION: Digital imaging in an autopsy environment can be both practical and cost-effective. It provides many advantages over traditional 35-mm photography and can be the first step toward numerous additional improved services.
Autopsy↗
Immunohistochemical distinction of ocular sebaceous carcinoma from basal cell and squamous cell carcinoma.
BACKGROUND: Diagnosis of sebaceous carcinoma of the periorbital region is often delayed. Clinically, this lesion can mimic several inflammatory disorders. Histopathologically, it can mimic either squamous cell or basal cell carcinoma. OBJECTIVE: To identify an immunohistochemical approach to assist in the diagnosis of periorbital sebaceous carcinoma. METHOD: The immunohistochemical profiles of several cases of periorbital sebaceous, basal cell, and squamous cell carcinoma were examined. RESULTS: Although at least focal epithelial membrane antigen (EMA) staining can effectively distinguish sebaceous carcinoma (10 of 11 were positive) from basal cell carcinoma (1 of 16 were positive), most squamous cell carcinomas examined were also focally EMA positive (11 of 14). However, Cam 5.2 reactivity was seen in most sebaceous carcinomas (8 of 11) but no squamous cell carcinomas (0 of 14). In addition, at least focal BRST-1 reactivity was also seen in most sebaceous carcinomas (7 of 11) but no basal cell carcinomas (0 of 16). CONCLUSIONS: Periorbital sebaceous, basal cell, and squamous cell carcinomas have different immunohistochemical staining profiles; a panel of commonly available antibodies, including anti-EMA, BRST-1, and Cam 5.2, may help distinguish these diseases from each other when that distinction cannot be clearly made by light microscopy alone.
Adenocarcinoma, Sebaceous↗
Exclusion of the stomatin, alpha-adducin and beta-adducin loci in a large kindred with dehydrated hereditary stomatocytosis.
Defects in stomatin, alpha-adducin, and beta-adducin have been implicated in erythrocyte disorders of cation permeability. We performed linkage analysis of the genetic loci for these proteins in a large kindred with xerocytosis (dehydrated hereditary stomatocytosis). Using polymerase chain reaction-based genotyping techniques, all three loci are excluded as disease gene candidates.
Alleles↗
Pigmented villonodular synovitis of the temporomandibular joint: diagnostic imaging and endovascular therapeutic embolization of a rare head and neck tumor.
We report a case of pigmented villonodular synovitis involving the temporomandibular joint that presented as a rapidly growing tumor with extension through the skull base into the middle cranial fossa. The case is of interest not only because of the unusual extensive infiltration of this tumor but also because of the role modern diagnostic imaging and endovascular therapeutic techniques played in its diagnosis and management.
Adult↗
Utilization of an 86 bp exon generates a novel adducin isoform (beta 4) lacking the MARCKS homology domain.
A novel isoform of beta-adducin has been amplified and characterized from a human bone marrow cDNA library (GenBank #U43959). This isoform arises from the insertion of an 86 bp alternatively spliced and previously unrecognized exon (now termed exon 15) within codon 581 of the human red blood cell beta-adducin sequence. This results in an insertion of 28 novel amino acids. The remainder of the red cell beta-adducin mRNA is then translated in a different reading frame, adding an additional 35 novel amino acids prior to the stop codon. This new isoform, thus, replaces beta 1-adducin sequence after residue 580 with a total of 63 new amino acids. Sequences from genomic clones of the human beta-adducin gene show that this alternate exon is flanked by splice consensus sequences and is appropriately located in the genomic map between exons encoding up-stream and down-stream sequences, thus defining a new exon. The COOH-terminus of this new isoform, which we designate beta 4, lacks a 22 amino acid lysine-rich sequence common to both the human red cell alpha- and beta-adducin subunits and homologous to a highly conserved region in MARCKS, a filamentous actin-cross linking protein regulated by protein kinase C and calcium/calmodulin. beta 4-adducin preserves a previously identified calmodulin binding domain. PCR analysis indicates that this new beta-adducin isoform is expressed in fetal brain and liver, bone marrow, and NT-2 (neuroepithelial) cells, but is not detected in several other tissues. We anticipate that this new beta 4 isoform of beta-adducin will display unique and tissue-specific functional properties.
Alternative Splicing↗
Neuronal loss from the subthalamic nuclei in a patient with progressive chorea.
We present a case of an 80-year-old man who developed a seizure disorder at age 66 and was treated with chronic phenytoin. In the last 3 years of his life, he developed multiple neurological deficits, including bilateral chorea, ataxic gait, sensory neuropathy, and progressive dementia. After death from pneumonia, autopsy examination of the patient's brain was most remarkable for a selective loss of neurons from both subthalamic nuclei and Purkinje cell loss in the cerebellum. This pattern of injury is consistent with a toxic process and does not fit previously characterized pathological syndromes known to be associated with movement disorders or dementia or both. Phenytoin has been shown to cause choreiform movements, peripheral neuropathy, and cognitive decline in some patients, but the pathological basis for these changes has not been elucidated. The patient's chorea was very likely the result of neuronal loss in the subthalamic nuclei, but causes for his dementia and neuropathy were not found. The pathological findings may represent either an unusual form of chronic phenytoin toxicity or a previously undescribed primary degenerative brain syndrome.
Aged↗
Reduced alpha-catenin and E-cadherin expression in breast cancer.
BACKGROUND: The expression of the homotypic cell adhesion protein, E-cadherin, is reduced in many types of cancer. The loss of this protein may be associated with metastasis because alteration of its function is required for invasion in vitro, and decreased expression has been associated with more aggressive tumor behavior in vivo. It is likely that the loss of downstream effector elements in the cadherin adhesion cascade may also disrupt cell-cell interactions and thereby promote invasion, but direct evidence for this has been lacking. One such effector element is alpha-catenin, a cytoplasmic protein related to vinculin that is associated in vivo with E-cadherin. EXPERIMENTAL DESIGN: In the present study, antibodies prepared to recombinant human alpha-catenin and recombinant human E-cadherin have been used to explore by immunocytochemistry the steady state levels of these proteins in a series of 26 cancers of the breast. RESULTS: The expression of alpha-catenin was reduced or lost more frequently (81% of cases) than was the expression of E-cadherin (63% of cases). Cases with absent E-cadherin expression uniformly lacked alpha-catenin. Eight of the 26 patients (31%) had known metastatic disease at the time of biopsy; yet, all patients with normal alpha-catenin staining in their tumors were free of known metastatic disease (four patients). CONCLUSIONS: Together with previous data on E-cadherin, these results suggest that reduced steady state levels of alpha-catenin may be a sensitive marker for disturbances in the adhesive function of the junctional complex and suggest that failure of at least one component of the cadherin-mediated cell-cell adhesion cascade is a common feature of breast, and presumably other, epithelial tumors.
Adult↗
Cardiogenic shock due to progression of cutaneous T-cell lymphoma.
We describe a patient with progressive cutaneous T-cell lymphoma (CTCL) and development of subacute cardiac failure. Symptomatic lymphomatous involvement of the heart may be more common in patients with CTCL than in other lymphomas because the former is more likely to be associated with circulating tumor cells and hematogenous spread to the myocardium. No single symptom or sign is highly predictive of cardiac involvement, but unexplained tachyarrhythmias, conduction disturbances, low voltage on ECG, and unexplained cardiac enlargement should arouse clinical suspicion. Although echocardiography may be helpful in suggesting cardiac involvement, endomyocardial biopsy should be considered in patients with a reasonable chance of responding to chemotherapy or radiation.
Electrocardiography↗
Acanthamoeba myosin-II minifilaments assemble on a millisecond time scale with rate constants greater than those expected for a diffusion limited reaction.
We have shown previously that Acanthamoeba myosin-II minifilaments assemble by three successive dimerization steps, forming, progressively, monomers, antiparallel dimers, antiparallel tetramers, and finally the full size octameric minifilament (Sinard, J. H., Stafford, W. F., and Pollard, T. D. (1989) J. Cell Biol. 109, 1537-1548). In the current study, we investigate the kinetics of the assembly of these minifilaments, initiating assembly by the rapid dilution of salt in a stopped-flow light scattering apparatus. The majority of the reaction is completed within 50 ms and is greater than 90% completed within 1 s. Assembly data over a greater than 6-fold myosin concentration range can be fit using the successive dimerization mechanism with a single set of rate constants. Second order rate constants for the initial steps in the assembly reaction exceed 10(8) M-1 s-1, and equilibrium dissociation constants predict a very low critical concentration, consistent with previous data. Other possible assembly mechanisms do not adequately fit all of the available data. Filament disassembly at 300 mM KCl is even more rapid, and there is both an increase in the dissociation rate constants and a decrease in the association rate constants with increasing KCl. Aggregation of minifilaments induced by Mg2+ is much slower and takes many minutes to reach equilibrium.
Acanthamoeba↗
Identification of functional regions on the tail of Acanthamoeba myosin-II using recombinant fusion proteins. II. Assembly properties of tails with NH2- and COOH-terminal deletions.
We used purified fusion proteins containing parts of the Acanthamoeba myosin-II tail to localize those regions of the tail responsible for each of the three steps in the successive dimerization mechanism (Sinard, J. H., W. F. Stafford, and T. D. Pollard. 1989. J. Cell Biol. 107:1537-1547) for Acanthamoeba myosin-II minifiliment assembly. Fusion proteins containing the terminal approximately 90% of the myosin-II tail assemble normally, but deletions within the last 100 amino acids of the tail sequence alter or prevent assembly. The first step in minifilament assembly, formation of antiparallel dimers, requires the COOH-terminal approximately 30 amino acids that are thought to form a nonhelical domain at the end of the coiled-coil. The second step, formation of antiparallel tetramers, requires the last approximately 40 residues in the coiled-coil. The final step, the association of two antiparallel tetramers to form the completed octameric minifilament, requires residues approximately 40-70 from the end of the coiled-coil. A region of the tail near the junction with the heads is important for tight packing of the tails in the minifilaments. Divalent cations induce the lateral aggregation of minifilaments formed from native myosin-II or fusion proteins containing a nonmyosin "head," but under the same conditions fusion proteins composed essentially only of myosin tail sequences with very little nonmyosin sequences form paracrystals. The region of the tail necessary for this paracrystal formation lies NH2-terminal to amino acid residue 1,468 in the native myosin-II sequence.
Acanthamoeba↗
Microinjection into Acanthamoeba castellanii of monoclonal antibodies to myosin-II slows but does not stop cell locomotion.
To study the in vivo role of myosin-II in Acanthamoeba castellanii, motile cells were microinjected with monoclonal antibodies raised against the myosin-II heavy chain. All injected cells underwent a transient shock response. It was found that although injection of buffer alone or of an endogenous Acanthamoeba protein decreased the motility of injected cells from 7 microns/min to approximately 3 microns/min, injection of monoclonal antibodies specific for myosin-II decreased motility further to approximately 0.8 micron/min. This effect was seen whether or not the monoclonal antibody to myosin-II inhibited the actomyosin-II MgATPase activity in vitro. Levels of antibody far in excess of endogenous myosin-II concentrations could not completely block amoeboid movement. The morphology of moving antimyosin-II-injected cells was unusual, suggesting a greater defect in the ability to retract the trailing edge of the cell rather than to extend the leading edge. Endosomes frequently disappeared from injected cells, and although buffer-injected cells rapidly recovered visible endosomes (50% recovery at 5 min), endosomes were not seen in antimyosin-II-injected cells until, on the average, approximately 50 min after injection. Injection of a nonspecific antibody or of a nonspecific exogenous protein (ovalbumin) also decreased the mobility of the injected cells beyond that of buffer-injected cells (to approximately 1 micron/min). These cells tended to recover endosomes more rapidly (approximately 25 min) than cells injected with antimyosin-II monoclonal antibodies. The inability of antibodies to myosin-II to inhibit completely any of the movements studied suggests that although myosin-II probably plays a role in these motilities, the cell either routinely uses or can draw upon another cytoplasmic motor to maintain locomotion, organelle movement, contractile vacuole activity, and endocytosis.
Acanthamoeba↗
Location of the head-tail junction of myosin.
The tails of double-headed myosin molecules consist of an alpha-helical/coiled-coil structure composed of two identical polypeptides with a heptad repeat of hydrophobic amino acids that starts immediately after a conserved proline near position 847. Both muscle and nonmuscle myosins have this heptad repeat and it has been assumed that proline 847 is physically located at the head-tail junction. We present two lines of evidence that this assumption is incorrect. First, we localized the binding sites of several monoclonal antibodies on Acanthamoeba myosin-II both physically, by electron microscopy, and chemically, with a series of truncated myosin-II peptides produced in bacteria. These data indicate that the head-tail junction is located near residue 900. Second, we compared the lengths of two truncated recombinant myosin-II tails with native myosin-II. The distances from the NH2 termini to the tips of these short tails confirms the rise per residue (0.148 nm/residue) and establishes that the 86-nm tail of myosin-II must start near residue 900. We propose that the first 53 residues of heptad repeat of Acanthamoeba myosin-II and other myosins are located in the heads and the proteolytic separation of S-1 from rod occurs within the heads.
Acanthamoeba↗
The effect of heavy chain phosphorylation and solution conditions on the assembly of Acanthamoeba myosin-II.
At low ionic strength, Acanthamoeba myosin-II polymerizes into bipolar minifilaments, consisting of eight molecules, that scatter about three times as much light as monomers. With this light scattering assay, we show that the critical concentration for assembly in 50-mM KCl is less than 5 nM. Phosphorylation of the myosin heavy chain over the range of 0.7 to 3.7 P per molecule has no effect on its KCl dependent assembly properties: the structure of the filaments, the extent of assembly, and the critical concentration for assembly are the same. Sucrose at a concentration above a few percent inhibits polymerization. Millimolar concentrations of MgCl2 induce the lateral aggregation of fully formed minifilaments into thick filaments. Compared with dephosphorylated minifilaments, minifilaments of phosphorylated myosin have a lower tendency to aggregate laterally and require higher concentrations of MgCl2 for maximal light scattering. Acidic pH also induces lateral aggregation, whereas basic pH leads to depolymerization of the myosin-II minifilaments. Under polymerizing conditions, millimolar concentrations of ATP only slightly decrease the light scattering of either phosphorylated or dephosphorylated myosin-II. Barring further modulation of assembly by unknown proteins, both phosphorylated and dephosphorylated myosin-II are expected to be in the form of minifilaments under the ionic conditions existing within Acanthamoeba.
Acanthamoeba↗
The mechanism of assembly of Acanthamoeba myosin-II minifilaments: minifilaments assemble by three successive dimerization steps.
We used 90 degrees light scattering, analytical ultracentrifugation, and electron microscopy to deduce that Acanthamoeba myosin-II minifilaments, composed of eight molecules each, assemble by a novel mechanism consisting of three successive dimerization steps rather than by the addition of monomers or parallel dimers to a nucleus. Above 200 mM KCl, Acanthamoeba myosin-II is monomeric. At low ionic strength (less than 100 mM KCl), myosin-II polymerizes into bipolar minifilaments. Between 100 and 200 mM KCl, plots of light scattering vs. myosin concentration all extrapolate to the origin but have slopes which decrease with increasing KCl. This indicates that structures intermediate in size between monomers and full length minifilaments are formed, and that the critical concentrations for assembly of these structures is very low. Analytical ultracentrifugation has confirmed that intermediate structures exist at these salt concentrations, and that they are in rapid equilibrium with each other. We believe these structures represent assembly intermediates and have used equilibrium analytical ultracentrifugation and electron microscopy to identify them. Polymerization begins with the formation of antiparallel dimers, with the two tails overlapping by approximately 15 nm. Two antiparallel dimers then associated with a 15-nm stagger to form an antiparallel tetramer. Finally, two tetramers associate with a 30-nm stagger to form the completed minifilament. At very low ionic strengths, the last step in the assembly mechanism is largely reversed and antiparallel tetramers are the predominant species. Alkaline pH, which can also induce minifilament disassembly, produces the same assembly intermediates as are found for salt induced disassembly.
Acanthamoeba↗