Biomedical subjects
Publications and source records attributed to L T Chow.
The postmortem examination of a 59-year-old man who had suffered from acute abdominal pain for two days revealed that the cause of death was peritonitis secondary to perforated peptic ulceration. During examination, it was found that the urinary tract was abnormal. Here, we report a postmortem finding of aplastic renal dysplasia of the kidney and malformation of the urinary tract.
Prolyl 4-hydroxylase, an alpha 2 beta 2 tetramer, plays a central role in collagen synthesis as it catalyzes the formation of 4-hydroxyproline residues by the hydroxylation of proline in X-Pro-Gly sequences. We report here that the human gene for the catalytically important alpha subunit is more than 69 kilobase pairs and consists of 16 exons. The exons that encode solely protein sequences vary from 54 to 240 base pairs (bp), and the introns vary from 750 to more than 16,000 bp. The 133 bp of 5'-untranslated sequences of the mRNA are coded by two exons, and these sequences contain inverted repeats with a potential for stem-loop formation, which may be involved in translational control of the expression of this gene. The 5'-flanking region contains a TATa motif at -29 relative to the major transcription site but no CCAAT motif. The 5'-flanking region and the downstream sequences contain several motifs that may act as binding sites for various transcription factors. Evidence has previously been reported for a mutually exclusive alternative splicing of RNA transcripts of this gene. The present data indicate that the mutually exclusive sequences found in the mRNAs are coded by two consecutive, homologous 71-bp exons 9 and 10. These exons are identical in their first 5 bp and the overall identity between them is 61% at the nucleotide level and 58% at the level of the coded amino acids. Both types of mRNA were found to be expressed in all of the tissues studied, but in some tissues the type coding for exon 9 or 10 sequences was more abundant than the other type.
We have established the first homologous cell-free DNA replication system for a papillomavirus. The replication of the human papillomavirus type 11 (HPV-11) origin was achieved by using human 293 cell extracts supplemented with the HPV-11 E1 and E2 proteins purified from insect cells infected with recombinant baculoviruses. Efficient replication depends on the HPV-11 origin, the HPV-11 E1 and E2 proteins, as well as human DNA polymerase alpha, delta, replication protein A, topoisomerase I, and topoisomerase II. High concentrations of E1 protein also promoted a low level of origin-independent replication which was suppressed by the addition of the E2 protein, whereas E2 protein stimulated origin-dependent replication. We also show that an intact E2 protein binding site was absolutely necessary for origin activity, as a strong HPV-11 origin was rendered inactive when one half-site of each of the three E2 binding sites was mutated. In contrast, there was only a relatively small reduction in this mutant origin activity when the cell extracts were supplemented with the bovine papillomavirus type 1 (BPV-1) proteins. These results suggest that the HPV-11 E2 protein plays a primary role in HPV origin recognition. Furthermore, unlike transient replication in which HPV-11 and BPV-1 viral proteins promote efficient replication of homologous and heterologous origins, efficient cell-free replication took place only with the homologous combinations.
The expression of proliferating cell nuclear antigen (PCNA) was studied in human papillomavirus (HPV)-infected, benign and malignant lesions of the genital tract and larynx using immunocytochemical staining of formalin-fixed clinical specimens. We observed the induction of PCNA in squamous carcinomas and adenocarcinomas, as has been demonstrated with other malignancies. In addition, the differentiated keratinocytes of the upper spinous cells and granulocytes in condylomata acuminata and low-grade intraepithelial neoplasias showed a consistent induction of PCNA compared with the normal squamous epithelium, in which only some of the parabasal and basal cells were positive. This reactivation of PCNA synthesis correlated with the presence of high copy numbers of HPV DNA and was independent of the oncogenic risk potential of the infecting HPV genotype. We postulate that HPV gene products induce the expression of PCNA and other components of the host DNA replication machinery in differentiated cells of squamous lesions to facilitate vegetative viral replication.
Kimura's disease is a chronic inflammatory disorder of unknown etiology, presenting usually as painless subcutaneous swellings in the head and neck region or in the salivary glands. The cytologic features of fine-needle aspirates of eight cases of Kimura's disease were studied with reference to the histologic appearance of the subsequent surgical specimens. In the cytologic smears, the prominent feature was the presence of significant numbers of eosinophils in a background of lymphoid cells. Fragments of collagenous tissue and Warthin-Finkeldey polykaryocytes occasionally were seen. In the cell block, vascular proliferation and fibrosis were useful features, providing further support to the diagnosis. The constellation of these features is characteristic of Kimura's disease and should suggest this diagnosis in the appropriate clinical setting. For initial diagnosis, excisional biopsy is important for the exclusion of malignant lymphoma, histiocytosis X, angiolymphoid hyperplasia with eosinophilia and other reactive lymphadenopathies. Nonetheless, fine-needle aspiration cytology may be valuable in the diagnosis of recurrent lesions of Kimura's disease and may spare the patient from repeated biopsies.
The ability to introduce DNA into mammalian cells has provided a powerful means to examine the regulation of gene expression and the function of gene products. However, the most commonly used techniques for DNA transfection are not always suitable for primary cells. Primary human keratinocytes are particularly stringent in their growth requirements and are also very refractory to transfection, rendering transient gene expression studies difficult. We have investigated the ability of several polycationic lipids to promote DNA uptake into human epidermal keratinocytes, as monitored with the bacterial beta-galactosidase reporter gene. We report that the cationic lipopolyamine dipalmitoyl phosphatidylethanolamine spermine as well as another procedure using Polybrene can achieve a 20% to 30% transfection efficiency, superior to any other agent tested on these cells. Gene transfer was accomplished by a 3-h exposure of monolayer cells to DNA complexes formed with either reagent by simple mixing in a serum-free medium, followed by a brief osmotic shock with glycerol. Neither DNA carrier showed any toxicity at the effective concentrations nor interfered with cell attachment, growth or differentiation. The use of a fully biodegradable lipopolyamine as DNA carrier should make it possible to extend this transfection method to gene transfer for in vivo therapeutic applications.
The E6 promoter of human papillomaviruses (HPVs) trophic for epithelia for the lower genital tract and the upper respiratory tract is regulated in vitro by homologous and heterologous papillomaviral E2 proteins that bind to a consensus responsive sequence (E2-RS) ACCN6GGT. When HPV type 11 (HPV-11) expression is examined in epithelial cell lines, the HPV-11 E2-C protein, which lacks the amino-terminal transactivating domain of the full-length E2 protein, invariably represses the homologous viral E6 promoter. In contrast, when the novel constitutive enhancer (CE) CE II is deleted, not only is the basal promoter activity much reduced, it is further repressed by the intact HPV-11 E2 protein (M. T. Chin, T. R. Broker, and L. T. Chow, J. Virol. 63:2967-2976, 1989). Here, we demonstrated that, when expressed from a stronger surrogate promoter, the HPV-11 E2 protein represses the E6 promoter effectively, regardless of CE II. By performing systematic mutational analyses of the four highly conserved copies of the HPV-11 E2-RS and of the adjacent enhancer-promoter elements, we show that the furthest upstream, promoter-distal E2-RS copy 1 plays no apparent role in E6 promoter regulation. Repression by the homologous HPV-11 E2 proteins is mediated through each of the three promoter-proximal copies of the E2-RS, but the presence of CE II abrogates the full-length E2 protein repression exerted at E2-RS copy 2. Repression is alleviated when the two (for E2) or three (for E2-C) promoter-proximal copies of E2-RS are mutated. We specifically demonstrate that repression exerted at E2-RS 3 is due to preclusion of binding of the host transcription factor Sp1 or Sp1-like proteins to a nonconsensus sequence AGGAGG located 1 bp upstream of the tandem E2 protein binding sites 3 and 4. A 3-bp insertion between the adjacent Sp1 and E2-RS 3 sites permits both Sp1 and E2 proteins to bind, with a concomitant relief of E2-RS 3-mediated repression. Similar mutational analyses show that proteins that bind to the GT-1 motif near the upstream E2-RS 2 help abrogate repression by the E2 protein in the presence of CE II. The implications of these results with respect to the viral infectious cycle and during viral oncogenesis are discussed.
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An 88-year-old man presented symptoms and signs of ascending cholangitis and died 20 days after the onset of illness. Postmortem examination revealed a mucinous tumor arising from the head of the pancreas, encasing the common bile duct and invading the liver with multiple hepatic metastasis. The tumor showed a unique and uniform histological appearance, consisting of signet-ring neoplastic cells floating in mucin pools. The rapid clinical course and widespread hepatic metastasis of this patient suggest that this pure, signet-ring variant of mucinous adenocarcinoma of the pancreas might have a poorer prognosis.
A 49-year-old woman with chronic renal failure was given gemfibrozil for hyperlipidemia. She developed gemfibrozil-induced myositis which precipitated an acute compartment syndrome, necessitating emergency fasciotomy. The muscle biopsy showed prominent degeneration of the skeletal muscle fibers, associated with moderate chronic inflammatory infiltration. Electron microscopy revealed myofibrillary fragmentation and mitochondrial disorganization. The clinicopathologic features of gemfibrozil-induced myositis appear to be distinct from those of clofibrate-induced muscular syndrome. Extreme caution should be exercised in the use of gemfibrozil in patients with impaired renal function.
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OBJECTIVE: To present a case of pulmonary arteriovenous malformation (AVM) that showed progressive enlargement in a patient with concomitant rheumatic mitral stenosis. CLINICAL FEATURES: A 73-year-old woman first presented eight years ago because of an uncomplicated acute myocardial infarction. She was noted to have rheumatic mitral stenosis. Six years later, she developed symptomatic sick sinus syndrome for which a permanent pacemaker was implanted. Chest x-ray revealed new development of an opacity in the middle zone of the right lung. Two years later, she was admitted to hospital because of progressive dyspnoea. Chest x-ray showed further increase in size of the right middle zone opacity. OUTCOME: She died of cardiopulmonary failure on the 18th day in hospital. Autopsy revealed a pulmonary AVM, replacing the entire right middle lobe. CONCLUSION: The elevated pulmonary vascular resistance caused by mitral stenosis may have increased the preferential flow of blood through the AVM, leading to its rapid enlargement. Patients with concomitant pulmonary AVM and mitral stenosis need to be recognised, as close follow-up and early treatment may be desirable.
OBJECTIVE: To present a case of gemfibrozil-induced myositis which precipitated an acute compartment syndrome. CLINICAL FEATURES: A 49-year-old woman with chronic renal failure was given gemfibrozil for hyperlipidaemia. She developed myositis in the sixth week of therapy. Her symptoms initially persisted despite withdrawal of gemfibrozil and she developed an acute compartment syndrome. INTERVENTION AND OUTCOME: Emergency fasciotomy was performed. She gradually improved and complete recovery occurred eight weeks after cessation of gemfibrozil therapy. CONCLUSION: Gemfibrozil may induce a local myositis which, in our patient, precipitated an acute compartment syndrome. One should be alert to symptoms of possible drug-induced myositis in patients receiving gemfibrozil. Extreme caution should be exercised in its use in patients with impaired renal function.
A patient in whom a free-floating but immobile ball thrombus developed in the left atrium in association with mitral stenosis is reported. Certain unusual echocardiographic findings are described.
Localized amyloidosis of the larynx is usually described as a non-bleeding lesion. We report a patient with localized laryngeal amyloidosis who developed a massive upper respiratory tract haemorrhage and died. This potentially fatal complication of localized amyloidosis of the larynx merits recognition as the disease could be controlled in most instances by surgical excision of the amyloid deposit.
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Two patients with localized polyarteritis nodosa involving the breast are reported. They presented with tender breast masses which were excised because of suspicion of carcinoma. The pathological features were similar with vasculitis of medium-sized and small arteries. The medium-sized arteries showed intimal fibroplasia, fragmentation of internal elastic lamina and inflammatory cellular infiltration. Fibrinoid vasculitis involving small arteries indicated a diagnosis of polyarteritis nodosa rather than giant cell arteritis. Both patients remain well. The differential diagnosis of these two arteritides and of Wegener's granulomatosis involving the breast is reviewed.
Human papillomavirus (HPV) type 11 is strictly trophic for epithelial cells and induces benign condylomata of the external genitalia and also causes laryngeal papillomas. Primary keratinocytes are the appropriate hosts for studies of HPV gene regulation, but they are not frequently used, owing to difficulties in culturing and low transfection efficiencies. By modifying a Polybrene transfection procedure, we achieved consistently high transfection efficiencies in primary human foreskin keratinocytes and characterized the HPV type 11 enhancer in the context of the homologous E6 promoter. Contrary to previous studies with immortalized human cervical carcinoma C-33A cells, constitutive enhancer element II in the upstream regulatory region conferred no enhancer activity and did not abrogate repression by the homologous E2 protein. Rather, repression was strong, ranging from 5.6- to 20-fold for the various enhancer deletion mutations. By deletion analysis, a strong enhancer that included three nuclear factor 1 sites and one nuclear factor 1-associated factor-binding site was localized to a 45-bp region within constitutive enhancer element I, and it showed some degree of tissue specificity.