Cloning and promoter analysis of the human B-50/GAP-43 gene.

We here report isolation of exon 1 and analysis of the human B-50 promoter. A human genomic lambda EMBL3 library was screened with a homologous PCR probe. Two independent clones were analyzed and partially sequenced: They contained up to 5 kb sequence upstream of the translation start site and approx 13 kb of intron 1 sequence. There was a high degree of homology between the rat and the human gene with 100% homology from -504 to -427, with respect to the translation start codon. However, relatively long GT and GA repeats as seen in the rat gene were absent. Various promoter-reporter constructs, containing 5.0 to 0.12 kb of the upstream region, were transfected into undifferentiated and neuroectodermally differentiated P19-EC. Two promoter activities were found. The minimal fragment with promoter activity still responsive to differentiation was the 0.22 kb construct, similar to rat promoter P2. We conclude that the human B-50 gene is expressed in a similar way to the rat B-50 gene, based on the presence of two transcripts, the high degree of homology between the rat and the human sequence, and the two promoter activities found in P19-EC cells.

Cyclosporine: a review and its specific use in liver transplantation.

During the past decade, treatment with cyclosporine, an immunosuppressive agent, has contributed substantially to enhanced allograft and patient survival after liver transplantation. Currently, the 1-year survival rate after liver transplantation is more than 80% in major liver transplantation centers, in contrast with approximately 60% before the availability of cyclosporine. Its predominant immunologic effect is inhibition of lymphokine production and secretion by helper T cells. The use of cyclosporine, however, is associated with numerous adverse effects, the most important of which are nephrotoxicity, hypertension, neurotoxicity, opportunistic infections, and malignant lesions. Acute nephrotoxicity, hypertension, and neurotoxicity usually can be reversed by decreasing the dose of cyclosporine. Measurement of cyclosporine concentrations in the blood is essential for optimization of immunosuppressive therapy and prevention of toxicity.

Cyclosporine, low-density lipoprotein, and cholesterol.

Lipoproteins are known to be able to transport a variety of drugs. This report suggests that low-density lipoprotein not only functions as an important carrier of cyclosporine in plasma but also facilitates transport of cyclosporine across the cell membrane by means of the low-density lipoprotein receptor. Such a mechanism would explain (1) the similar tissue distribution of cyclosporine and the low-density lipoprotein receptor, (2) the increase in immunosuppression and toxicity with low total serum cholesterol levels, and (3) the relative absence of immunosuppression and toxicity with high levels of cyclosporine in the blood in patients with hypertriglyceridemia. In addition to receptor-mediated uptake, a disturbance of the blood-brain barrier is suggested as an explanation of the high frequency of cyclosporine-induced central nervous system toxicity after liver transplantation. Cyclosporine-induced inhibition of the mitochondrial steroid 26-hydroxylase, an enzyme involved in the formation of bile acids from cholesterol and deficient in patients with cerebrotendinous xanthomatosis, may cause or contribute to the observed central nervous system toxicity. It also may explain the similar clinical features of cyclosporine-induced central nervous system toxicity and cerebrotendinous xanthomatosis.

Dysarthria and apraxia of speech associated with FK-506 (tacrolimus).

The immunosuppressive agent FK-506 (tacrolimus) is one of the agents most commonly used to prevent rejection after liver transplantation. Neurologic toxicity related to FK-506 has been reported, including speech disorders; however, a detailed analysis of the speech disorder associated with use of FK-506 has not been presented. Herein we describe a patient who exhibited mutism, then severe apraxia of speech with a concomitant hypokinetic, spastic, and ataxic dysarthria after administration of FK-506. His residual mixed dysarthria, without radiographic evidence of a structural lesion, suggests dysfunction of one or more neurochemical systems. The pathophysiologic mechanisms underlying this intriguing entity remain obscure.

Unusual aspects of acute Q fever-associated hepatitis.

We describe a 37-year-old farmer with a 3-week history of fevers and hepatitis, in whom Q fever was diagnosed. The diagnosis was based on the findings of characteristic "ring" granulomas on a bone marrow biopsy specimen and confirmed by complement-fixation antibody tests to Coxiella burnetii. Unusual aspects of this case included (1) relatively low complement-fixation antibody titers, (2) prolonged prothrombin time, (3) false-positive results of a serologic test for the human immunodeficiency virus (HIV), and (4) ring granulomas that progressed to atypical granulomas in biopsy specimens.

Intracerebral hemorrhage in liver transplant recipients.

OBJECTIVE: To analyze possible causative mechanisms for intracranial hemorrhage after orthotopic liver transplantation. DESIGN: We conducted a retrospective survey of medical records and a case-control comparison in patients who had undergone liver transplantation during the period 1986 through 1992. MATERIAL AND METHODS: In a group of 8 patients with intracranial hemorrhage after orthotopic liver transplantation and a control series of 207 patients who had undergone liver transplantation but did not have intracranial hemorrhage, we summarized pertinent clinical and laboratory data and statistically analyzed potential risk factors for hemorrhage. RESULTS: In the eight study patients, intracerebral hematomas were located in the parietal or frontal lobe in six, the cerebellum in one, and the putamen in one. Autopsy demonstrated a Candida-associated mycotic aneurysm in one of the eight patients, and one had disseminated aspergillosis. No statistically significant differences in thrombocytopenia, hypertension, extracranial bleeding sites, or cyclosporine-related neurotoxicity were found when these patients were compared with the control series. Bacteremia or fungemia was found in five of the eight patients with intracerebral hemorrhages (62%) but in only 11% of the control group (P = 0.03; Fisher's exact test). CONCLUSION: Overwhelming infections, thrombocytopenia, or both may have a role in intracerebral hemorrhage after liver transplantation.

Angiotropic large cell lymphoma (intravascular lymphomatosis) occurring after follicular small cleaved cell lymphoma.

Angiotropic large cell lymphoma is a rare, aggressive type of malignant lymphoma that primarily involves intravascular spaces and most often has clinical manifestations in the skin and central nervous system. Virtually any organ can be affected, however, including the lymph nodes and spleen. Peripheral blood involvement is usually not detectable morphologically. Conventional lymphoma in association with this entity has also been described. Herein we present a case of angiotropic lymphoma of B-cell lineage that affected the liver and skin. Reanalysis of a lymph node specimen that had been excised 3 years previously demonstrated a follicular small cleaved cell lymphoma. To our knowledge, this is the first reported case of possible evolution of follicular lymphoma to large cell lymphoma of angiotropic type.

Improved diagnostic yield of endoscopic biliary brush cytology by digital image analysis.

OBJECTIVE: To evaluate the accuracy of digital image analysis (DIA) for distinguishing between benign and malignant strictures of the biliary tract. PATIENTS AND METHODS: Our pathology databank was used to identify all biliary brush cytology specimens obtained during endoscopic retrograde cholangiopancreatography between June 1997 and June 1999. Corresponding medical records were reviewed to determine whether patients had benign or malignant strictures. Strictures were further classified into benign strictures with negative routine cytology, malignant strictures with negative routine cytology, and malignant strictures with positive routine cytology. Papanicolaou-stained smears of available brush cytology specimens were destained and then restained with Feulgen dye. Nuclear images were quantified for DNA content without knowledge of stricture type. DNA histograms were generated and ploidy results compared with the class of stricture. RESULTS: We analyzed 27 specimens from 69 confirmed benign or malignant strictures. Assuming that the presence of any aneuploid cells indicated malignancy, the sensitivity of DIA was 85%. Furthermore, aneuploid cells were detected by DIA in 13 of 16 specimens in which routine cytology was unrevealing. CONCLUSION: Ploidy assessment by DIA has potential to enhance the sensitivity of diagnosing malignant strictures compared with routine cytology alone.

Hepatic lymphangiomatosis mimicking polycystic liver disease.

Hepatic lymphangiomatosis is a rare disorder characterized by cystic dilatation of the lymphatic vessels in the hepatic parenchyma. It can occur in the liver alone, in the liver and spleen, or in multiple organs. Clinically, diagnosis can be difficult because of the rarity and protean manifestations of this disorder. We describe a 53-year-old woman with hepatic lymphangiomatosis in whom polycystic liver disease had been previously diagnosed. In addition, we review 12 cases of hepatic, splenic, and hepatosplenic lymphangiomatosis with or without systemic lymphangiomatosis and discuss the differential diagnosis.

Structure of the human gene for the neural phosphoprotein B-50 (GAP-43).

The genomic DNA encoding the exons for the human neural phosphoprotein B-50 (GAP-43) was isolated using rat-based cDNA probes and oligonucleotides. Exons 2 and 3 were isolated from a genomic library, exon 1 was amplified by PCR on total genomic DNA. The gene consists of 3 exons and 2 large introns. The first exon encodes the N-terminal 10 amino acids of B-50 involved in membrane association of the protein. Exon 2 encodes the main part of the protein with the sites for protein kinase C-mediated phosphorylation and calmodulin binding, and includes a 10 amino acid residue insert not found in rodents. Exon 3 encodes the last 29 amino acid residues. The reported sequence extends the known cDNA structure to both the 5' and 3' ends. The 358 bp region upstream of the translational initiation codon, containing the main transcription starts, is purine-rich and does not include TATA or GC boxes. At the 3' end potential polyadenylation signals were found 510 bp and 584 bp downstream of the stopcodon in exon 3. The 5' end of the mRNA is heterogeneous in length, with primer extension products corresponding to a 5' untranslated region of 159 and 343 bases. Northern hybridizations, however, indicate that the majority of B-50 mRNA has a shorter 5' untranslated region, as was reported for the rat (Schrama et al., Soc. Neurosci. Abstr., 18 (1992) 333.4). The structural organization of the human gene is similar to that described for the rat (Grabczyk et al., Eur. J. Neurosci. 2 (1990) 822-827), and both translated and untranslated regions show a high degree of sequence homology to the rat gene.

Characterization and growth regulation of a rat intrahepatic bile duct epithelial cell line under hormonally defined, serum-free conditions.

Bile duct epithelial cells, or cholangiocytes, proliferate in vivo under a number of pathologic (i.e., partial hepatectomy) and pathophysiologic (i.e., bile duct ligation, malignant transformation) conditions. However, little is known about the possible growth factors that modulate these proliferative responses, in part because an in vitro model to study proliferation of nontransformed, normal cholangiocytes is not available. We report here the development of a rat cholangiocyte cell line (MMRC, minimal media-requiring rat cholangiocytes) that grows under hormonally defined, serum-free conditions on plastic and maintains a cholangiocyte phenotype. Morphologic as well as functional studies indicate that the cell line is polarized and actively transports fluid and electrolytes in an apical to basolateral direction. MMRC, when cultured for 24 mo. and passaged 80 times, have not undergone malignant transformation, because the cell line failed to grow under anchorage-independent conditions or in nude mice. Cellular proliferation is accelerated 2-8-fold by insulin, insulin-like growth factor 1, epidermal growth factor, and hepatocyte growth factor, growth factors known to stimulate tyrosine kinase receptors, and inhibited 2-10-fold by TGFbeta and IL-2. Glyco-conjugates of primary (i.e., cholic and chenodeoxycholic acid) and secondary bile acids (i.e., deoxycholic and lithocholic acid) do not alter proliferation at low concentration (1 microM), but are toxic at higher concentration (10 microM). In summary, we have developed and characterized a cholangiocyte cell line derived from normal rat liver, which grows under hormonally defined, serum-free conditions, maintains a nonmalignant, cholangiocyte phenotype, displays morphologic and functional features of polarity, and alters its proliferation rate in response to a variety of growth factors.

Eosinophil-mediated bile duct stricture. A case studied by immunohistochemistry.

Major basic protein accounts for the majority of the protein within the eosinophilic granule. Utilizing immunohistochemical staining for major basic protein, we have demonstrated the dominant role of the eosinophil in a reversible bile duct stricture.

World-Wide Web-based graphical user interfaces for laboratory data.

OBJECTIVES: Electronic medical record systems permit collection of large amounts of medical information. Usually, information is presented in a fixed format, either as text or tables. Health care providers have to navigate this fixed format in order to find information useful for a specific patient-provider interaction. The main objective of this work was to allow the provider immediate access to specific laboratory information through the development of a highly customizable, graphical user interface to the Mayo Clinic laboratory information system. RESULTS: Here we describe this platform-independent, World-Wide-Web-based graphical user interface that allows the provider to see all or a predetermined panel of essential laboratory data in graphical format. Advantages include availability at internet-based workstations, immediate recognition of trends over time, ability to zoom in and out of specific periods of time, and detailed analysis of patient values in relationship to normal values. CONCLUSIONS: Web browser-based user interface allowing graphical display of laboratory data using Java technology was described. The connection to the Mayo Clinic laboratory information system combines cross-platform support for use on virtually any networked machine, interaction through a Web browser for ease of use, and a combination of the Perl and Java languages for powerful data processing and interactivity.

Near real-time retroflexion detection in colonoscopy.

Colonoscopy is the most popular screening tool for colorectal cancer. Recent studies reported that retroflexion during colonoscopy helped to detect more polyps. Retroflexion is an endoscope maneuver that enables visualization of internal mucosa along the shaft of the endoscope, enabling visualization of the mucosa area that is difficult to see with typical forward viewing. This paper describes our new method that detects the retroflexion during colonoscopy. We propose region shape and location (RSL) features and edgeless edge cross-section profile (ECSP) features that encapsulate important properties of endoscope appearance and edge information during retroflexion. Our experimental results on 50 colonoscopy test videos show that a simple ensemble classifier using both ECSP and RSL features can effectively identify retroflexion in terms of analysis time and detection rate.

Novel information theory based method for superimposition of lateral head radiographs and cone beam computed tomography images.

OBJECTIVES: The aim was to introduce a novel alignment criterion, focus mutual information (FMI), for the superimposition of lateral cephalometric radiographs and three dimensional (3D) cone beam computed images as well as the assessment of the alignment characteristics of the new method and comparison of the novel methodology with the region of interest (ROI) approach. METHODS: Implementation of a FMI criterion-based methodology that only requires the approximate indication of stable structures in one single image. The robustness of the method was first addressed in a phantom experiment comparing the new technique with a ROI approach. Two consecutive cephalometric radiographs were then obtained, one before and one after functional twin block application. These images were then superimposed using alignment by FMI where the following were focused on, in several ways: (1) cranial base and acoustic meatus, (2) palatal plane and (3) mandibular symphysis. The superimposed images were subtracted and coloured. The applicability to cone beam CT (CBCT) is illustrated by the alignment of CBCT images acquired before and after craniofacial surgery. RESULTS: The phantom experiment clearly shows superior alignment when compared to the ROI approach (Wilcoxon n = 17, Z = -3.290, and P = 0.001), and robustness with respect to the choice of parameters (one-sample t-test n = 50, t = -12.355, and P = 0.000). The treatment effects are revealed clearly in the subtraction image of well-aligned cephalometric radiographs. The colouring scheme of the subtraction image emphasises the areas of change and visualizes the remodelling of the soft tissue. CONCLUSIONS: FMI allows for cephalometry without tracing, it avoids the error inherent to the use of landmarks and the interaction of the practitioner is kept to a minimum. The robustness to focal distribution variations limits the influence of possible examiner inaccuracy.