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Biomedical subjects

M Landon

Publications and source records attributed to M Landon.

At least 19 recordsLinked to original sources
PubMed1992

The cloning and nucleotide sequence of the serine protease gene (aspA) of Aeromonas salmonicida ssp. salmonicida.

The gene for Aeromonas salmonicida serine protease has been cloned into phagemid pTZ18R in two restriction fragments, 2.0-kb PstI and 2.3-kb KpnI, of genomic DNA. The nucleotide sequences of the two fragments have been determined, in both directions, after subcloning, by double-stranded sequencing of nested deletions. An open reading frame of 1863 bp translated into a sequence of 621 amino acids, a 24-amino acid signal peptide and a 597-amino acid mature enzyme of molecular mass 64,173 Da. The consensus sequence, NGTS, of a serine protease substrate primary binding site was identified and a putative ribosome-binding site GGAG occurred 6 bp upstream of the ATG initiation codon.

Aeromonas↗
PubMed1992

Protein processing in lysosomes: the new therapeutic target in neurodegenerative disease.

A little recognised feature of neurons is their large complement of lysosomes. Studies of the accumulation of the abnormal isoform of the prion protein (PrPSC) in the prion encephalopathies and the formation of beta/A4 protein from its precursor in Alzheimer's disease suggest that generation of these key proteins takes place in lysosome-related organelles. The release of hydrolytic enzymes from lysosomes may be a primary cause of neuronal damage. Although molecular genetic approaches have identified protein mutations central to the main neurodegenerative disease, cell biological observations are now beginning to unravel the intracellular pathways involved in the molecular pathogenesis of neurodegeneration: as a result, it is now appropriate to consider therapeutic manipulation of the lysosomal system as an approach to treatment.

Animals↗
PubMed1992

alpha B crystallin expression in non-lenticular tissues and selective presence in ubiquitinated inclusion bodies in human disease.

alpha B crystallin is a lens protein which has homology with the small heat-shock proteins and is also expressed in non-lenticular tissues. Polyclonal antibodies have been raised to a synthetic peptide corresponding to residues 1-10 of alpha B crystallin. The antiserum detects a 20 kDa polypeptide on nitrocellulose replicas after polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate of extracts of heart muscle known to be rich in alpha B crystallin. Staining of normal human tissues reveals immunoreactivity of lens capsular epithelium, skeletal muscle, cardiac muscle, smooth muscle, renal tubular epithelium, Schwann cells, and glial cells, as has been described by other workers. In addition, positive staining of normal thyroid epithelium, colonic epithelium, and stratified squamous epithelium was seen. Tissues known to contain ubiquitinated inclusion bodies were immunostained with the anti-alpha B-crystallin antiserum. Staining of cortical Lewy bodies, astrocytic Rosenthal fibres, and hepatic Mallory bodies was seen, but only a proportion of inclusions were positive. Neurones containing the ubiquitinated inclusions of Alzheimer's disease were only very rarely immunostained and the ubiquitinated inclusions of motor neurone disease were not detected by the antiserum. Reactive astrocytes in cerebral tissues were strongly immunostained. The results suggest that alpha B crystallin is involved in the formation of ubiquitinated inclusion bodies that have associated intermediate filaments and support previous observations on the localization of a brain-specific ubiquitin carboxy-terminal hydrolase which similarly divides ubiquitinated filamentous inclusions in the central nervous system into two main groups.

Alzheimer Disease↗
PubMed1992

Lysosomes as key organelles in the pathogenesis of prion encephalopathies.

The causation, structural origin, and mechanism of formation of spongiform lesions in transmissible encephalopathies are unknown. We have used immunogold electron microscopy to locate ubiquitin conjugates, hsp 70, and beta-glucuronidase (markers of the lysosomal compartment) and prion protein (PrP) in both control and scrapie-infected mouse brain. In scrapie-infected brain, lysosomes and lysosome-related structures (multivesicular and tubulovesicular dense bodies) are present in abnormally high numbers in neuronal cell processes. These structures contain PrP, together with the lysosomal markers ubiquitin conjugates, hsp 70, and beta-glucuronidase, which could also be identified spilling from tubulovesicular dense bodies into areas of early rarefaction in neuronal processes; we suggest that these areas of rarefaction are the precursor lesions of spongiform change. We advance the hypothesis that spongiform change is brought about by cytoskeletal disruption in neuronal processes caused by liberation of hydrolytic enzymes from lysosomes overloaded with the abnormal isoform of PrP (PrPsc). We suggest that the lysosomal system is probably acting as the bioreactor for processing of normal PrP to the abnormal isoform. The continuous production of increasing quantities of abnormal PrPsc in lysosome-related bodies will eventually cause disruption of the lysosomal membrane with destruction of the neuronal cytoskeleton and the initiation of vacuolation. Later, death of the cell will be associated with release of the PrPsc isoform into the extracellular environment. Repeated rounds of phagocytosis, lysosomal biogenesis of PrPsc, lysosomal membrane rupture, hydrolytic enzyme release, and neuronal lysis will lead to an exponential increase in cell damage and cell death.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

Immunoreactivity to ubiquitin-protein conjugates is present early in the disease process in the brains of scrapie-infected mice.

Brains from mice infected with either the 87V or the ME7 strains of mouse-passaged sheep scrapie were taken at stages during the disease process and immunostained to show the localization of ubiquitin-protein conjugates. In both models, conjugates were seen as fine, dot-like structures; as coarser, granular lesions within or adjacent to neurones; and in areas surrounding plaques. The dot-like structures were visible at 28 days post-ME7 infection and at 55 days in 87V-infected mice. In both models, the extent of immunoreactive changes increased as the disease progressed and terminal infection was as described earlier by us (Lowe et al., J. Pathol 1990; 162: 61-66). The patterns of development of these features were distinctive in two ways: progression from region to region was observable and the density of the pathological lesions grew exponentially as the clinical symptoms appeared. The earliest pathological dot-like structures corresponded temporally with the earliest detection of PrPSC by Western blotting, and immunogold electron microscopic investigation of the dot-like lesions indicated that they were the multi-vesicular, lysosome-related, dense bodies that we have described previously in terminal disease (Laszlo et al., J Pathol 1992; 166: 333-341). Until now, ubiquitin-protein conjugates were seen mainly in inclusion bodies associated with the terminal stages of a range of human degenerative diseases. This study establishes that ubiquitin-protein conjugates accumulate in lysosome-related bodies very early and appear to be intimately related to the pathological processes in the animal disorders that we have studied.

Animals↗
PubMed1992

Ballooned neurons in several neurodegenerative diseases and stroke contain alpha B crystallin.

alpha B crystallin is a protein which has homology with the small cell stress proteins. A characterized antibody to residues 1-10 of alpha B crystallin was used to immunostain tissues containing ballooned (chromatolytic, achromasic) neurons. The tissues included two cases of classical Pick's disease, one case of dementia with swollen achromasic neurons in the cortex, two cases of Alzheimer's disease with large numbers of ballooned neurons, two cases of motor neuron disease, four cases of cortico-basal degeneration, and four cases with areas of brain showing swollen neurons adjacent to recent cerebral infarcts. The anti-alpha B crystallin showed strong diffuse cytoplasmic immunoreactivity of swollen cortical neurons in all the diseases. Astrocytes and oligodendroglial cells were also stained in normal tissues as previously described. Weak diffuse immunoreactivity with an antibody to ubiquitin-conjugates was also seen in the swollen neurons from cases of neurodegenerative disease but not following infarction. Ballooned neurons have been shown to contain phosphorylated neurofilament epitopes not normally present in the perikaryonal region. The presence of alpha B crystallin in ballooned neurons, together with previous data which also indicate its close association with intermediate filaments, suggest that alpha B crystallin may be involved in aggregation and remodelling of neurofilaments in disease. The presence of alpha B crystallin in neurons at the edge of areas of cerebral infarction is likely to reflect cells which are regenerating following damage; its detection may therefore be a marker for such cells. On a practical level, the antibody greatly facilitates the localization of such abnormal neurons in diagnostic histology.

Alzheimer Disease↗
PubMed1992

Buccal pulse oximeter is more accurate than finger pulse oximeter in measuring oxygen saturation.

Although there have been several anecdotal reports of the use of buccal pulse oximeter monitoring (Spo2) when digital Spo2 monitoring cannot be used, there have been no objective evaluations of the accuracy of buccal Spo2 monitoring. The purpose of this study was to systematically compare buccal Spo2 monitoring to both digital Spo2 and arterial O2 saturation monitoring (Sao2) in both generally anesthetized patients in the operating room (n = 31) and critically ill patients in the intensive care unit (n = 23). Buccal Spo2 probes were prepared by taping a malleable metal bar securely over the back of a Nellcor Oxisensor D-25 probe and bending the metal bar and buccal probe firmly around the corner of the patient's mouth. All buccal and finger Spo2 and Sao2 measurements were made simultaneously during hemodynamic stability. We found that buccal Spo2 was higher than finger Spo2 and agreed more closely with Sao2 for both patient groups (98.1% +/- 2.6%, 96.8% +/- 3.5%, 98.5% +/- 2.5%, respectively [mean +/- SD]). The operating room patients had higher buccal and finger Spo2 and Sao2 (99.3% +/- 1.5%, 98.9% +/- 1.4%, 99.5% +/- 0.7%, respectively) than the intensive care unit patients (96.4% +/- 2.9%, 94.1% +/- 3.5%, 96.6% +/- 3.5%, respectively). Although buccal Spo2 monitoring has several disadvantages (i.e., the probe requires preparation, can be more difficult to place, may be less readily accepted in awake patients, and is often mechanically dislodged during airway maneuvers), we conclude that buccal Spo2 monitoring is a more than adequate oximetry alternative when digital Spo2 monitoring is not an option (digits are unavailable or available digits are mechanically interfered with).

Blood Gas Analysis↗
PubMed1991

Ubiquitin in health and disease.

Studies in recent years have shown that ubiquitin has increasingly important functions in eukaryotic cells; roles which were previously not suspected in healthy and diseased cells. The interplay between molecular pathological and molecular cell biological findings has indicated that ubiquitin may be pivotal in the cell stress response in chronic degenerative and viral diseases. Furthermore, the studies have led to the notion that ubiquitination may not only serve as a signal for nonlysosomal protein degradation but may be a unifying covalent protein modification for the major intracellular protein catabolic systems; these can act to identify proteins for cytosolic proteinases or direct intact and fragmented proteins into the lysosome system for breakdown to amino acids. This unifying role could explain why ubiquitin is restricted to eukaryotic cells, which possess extensive endomembrane systems in addition to a nuclear envelope. Protein ubiquitination is a feature of most filamentous inclusions and certain other intracellular conglomerates that are found in some degenerative and viral diseases. The detection of ubiquitin-protein conjugates is not of great diagnostic importance in these diseases. Protein ubiquitination is not only essential for the normal physiological turnover of proteins but appears to have been adapted as part of an intracellular surveillance system that can be activated by altered, damaged, or foreign proteins and organelles. The purpose of this system is to isolate and eliminate these noxious structures from the cell: as a cytoprotective mechanism this appears to have evolved in the cell akin perhaps to an 'intracellular immune system'. Other heat shock proteins such as hsp 70 may be involved in this process. It is apparent that ubiquitin has a role in embryonic development. Protein ubiquitination is presumably involved in the reorganisation of cytoplasm that accompanies cell differentiation. Ubiquitin is also necessary for the gross intracellular degradative processes which are consequent upon programmed cell death. Cell elimination is of key importance for a number of developmental morphogenetic changes. An understanding of the molecular details of these processes will no doubt provide further insights into the wide ranging roles of ubiquitin in the life process. As it says in the book 'Ubiquitin'; there is no doubt that ubiquitin is a 'lucky' protein. It is lucky in many ways: lucky for scientific progress, lucky for biomedical scientists and lucky for life! If you have not already done so, why don't you get lucky and look for a role for ubiquitin in your experimental system. As Avram Hershko has said "there is plenty to go round"!

Animals↗
PubMed1991

Immunogold localisation of ubiquitin-protein conjugates in primary (azurophilic) granules of polymorphonuclear neutrophils.

Ubiquitin-protein conjugates are found in the primary (azurophilic) lysosome-related granules but not in the secondary (specific) granules in mature polymorphonuclear neutrophils prepared from bone marrow. This is the first reported demonstration of ubiquitin-protein conjugates in lysosome-related membrane-bound vesicles in granulocytes and complements our previous findings of ubiquitinated proteins in lysosomes of fibroblasts. The significance of the selective presence of conjugates in only one of the two main types of neutrophil granules remains to be elucidated but may relate to the presence of the complement of acid hydrolases, including proteases, in the azurophilic granules compared to the specific granules. Ubiquitin-protein conjugates may enter the primary granules during neutrophil maturation by an autophagic process or by a heterophagic process during the fusion of phagosomes with primary granules. Alternatively protein ubiquitination may be involved in granule biogenesis.

Animals↗
PubMed1991

The latent membrane protein-1 in Epstein-Barr virus-transformed lymphoblastoid cells is found with ubiquitin-protein conjugates and heat-shock protein 70 in lysosomes oriented around the microtubule organizing centre.

Immunofluorescence studies on Epstein-Barr virus (EBV)-transformed lymphoblastoid cells have previously shown that the latent membrane transforming protein (LMP-1) is found in patch-like inclusions which also immunostain for vimentin. We now show that EBV transformation causes a major reorganization of intermediate filaments, microtubules, mitochondria, and lysosomal elements, which generally become oriented around the microtubule organizing centre. Immunogold electron microscopy shows that LMP-1 is primarily concentrated in secondary lysosomes together with ubiquitin-protein conjugates and heat-shock protein 70. Intermediate filament inclusion formation with the above characteristics may be a general response triggered by other membrane glycoproteins; as seen, for example, in major human neurodegenerative diseases such as diffuse Lewy body disease.

Antigens, Viral↗
PubMed1991

Screening recommendations for gestational diabetes mellitus.

As a result of extensive experiences in multiple centers and a review of the current literature, we conclude that a plasma glucose level obtained 1 hour after a 50 gm oral glucose challenge is the "best" gestational diabetes mellitus screening test. This universal screening is performed at least once during pregnancy. The screening threshold should be no higher than 140 mg/dl, or an unacceptable loss in sensitivity occurs. Universal screening for gestational diabetes mellitus is justified by morbidity reduction, cost, and protocol simplicity and ease.

Blood Glucose↗
PubMed1991

Ubiquitin and the lysosome system: molecular immunopathology reveals the connection.

Considerable advances in our understanding of the molecular pathology of the major human neurodegenerative diseases have been made by the use of ubiquitin immunocytochemistry. The technique demonstrates that filamentous inclusions and vacuoles contain ubiquitin-protein conjugates. The molecular structure of the filaments and the morphological type of vacuoles is not completely understood but there is evidence that some of the filamentous inclusions contain intermediate filaments and the perinuclear distribution of the vacuoles resemble the distribution of intraneuronal lysosomes. Intermediate filaments and lysosomes are involved in the sequestration and degradation of viral membrane proteins in tissue culture cells. Immunogold electron microscopical and biochemical evidence indicates that ubiquitin-protein conjugates are normally considerably enriched in the lysosomes of fibroblasts relative to all other organelles. Immunogold electron microscopy shows a similar enrichment of ubiquitin-protein conjugates in the dense lysosomes of granulocytic precursor cells in the bone marrow. Filamentous inclusions showing several of the features seen in inclusions in the neurodegenerative diseases are seen in Epstein-Barr virus transformed lymphoblastoid cells. Immunohistochemistry shows that the inclusions contain vimentin intermediate filaments, the latent membrane transforming protein of the virus, ubiquitin-protein conjugates, and a heat-shock protein 70 (hsp 70). Immunohistochemistry and immunogold electron microscopy demonstrate that the latent membrane protein, ubiquitin-protein conjugates and hsp 70 are in lysosomes entwined in an intermediate filament cage centred on the microtubule organising centre. The implications of the combined observations for our understanding of the cell stress response in degenerating neurones and in virally infected cells are discussed.

Heat-Shock Proteins↗
PubMed1991

The role of protein ubiquitination in neurodegenerative disease.

Ubiquitin immunocytochemistry with an antiserum which reacts with ubiquitin-protein conjugates demonstrates the presence of ubiquitinated proteins in filamentous inclusions found in neurones in the major human neurodegenerative diseases, i.e. Alzheimer's disease, diffuse Lewy body disease, motor neurone disease. Ubiquitin immunohistochemistry has revolutionized the neuropathological diagnosis of dementia showing that diffuse Lewy body disease is not, as previously supposed, a rare cause of dementia. The filamentous inclusions in neurones in the human neurodegenerative diseases can be divided into at least two types based on recent immunocytochemical studies. We have shown that a ubiquitin-carboxyl terminal hydrolase is present in Lewy bodies but not in neurofibrillary tangles in Alzheimer's disease. This observation is significant since it indicates that molecular pathological mechanisms in neurones in diffuse Lewy body disease are fundamentally different to Alzheimer's disease. Ubiquitin-protein conjugates are also found in vacuoles in areas of granulovacuolar degeneration in hippocampal neurones in Alzheimer's disease and in granulovacuoles in neurones of scrapie infected mouse brain. These locations suggest that ubiquitinated protein are present in the lysosome-related system of neurones. We have recently shown that ubiquitin-protein conjugates are indeed enriched some 12-fold in the lysosomes of normal fibroblasts and lymphocytes.

Brain Diseases↗