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P D Hart

Publications and source records attributed to P D Hart.

At least 19 recordsLinked to original sources
PubMed1992

Amalgamation of two endocytic probe techniques: fluoresceinated cationized ferritin can show up, sequentially, selected structures, first in living cells and then by electron microscopy.

Fluorescein isothiocyanate-cationized ferritin (FITC-CF) has hitherto been used mainly to identify structures in living cells by light microscopy, by virtue of its fluorescent properties. We show here that this conjugate can be used, after immediate fixation of the same cell sample and preparation of thin sections, to recognise the same structures, by virtue of the ferritin's electron opacity. The conjugate should thus have a new use as a single-application, dual-purpose probe, e.g. in endocytic studies. The procedure may have advantages over similar dual-purpose probes in not requiring staining or special treatment.

Animals↗
PubMed1991

Ammonium chloride, an inhibitor of phagosome-lysosome fusion in macrophages, concurrently induces phagosome-endosome fusion, and opens a novel pathway: studies of a pathogenic mycobacterium and a nonpathogenic yeast.

The weak base ammonium chloride has been previously reported to inhibit lysosomal movements and phagosome-lysosome (Ph-L) fusion in cultured mouse macrophages (M phi), thus reducing delivery, to an intraphagosomal infection, of endocytosed solutes that have concentrated in secondary lysosomes. We have now addressed the question, whether NH4Cl might affect any direct interaction (if it exists) between such infection phagosomes and earlier, nonlysosomal compartments of the endocytic pathway, i.e., solute-containing endosomes. The phagosomes studied were formed after ingestion of the mouse pathogen Mycobacterium microti and the nonpathogenic yeast Saccharomyces cerevisiae; and the endosomes were formed after nonreceptor-mediated endocytosis of electronopaque and fluorescent soluble markers. By electron microscopy, survey of the cell profiles of M phi that had been treated with 10 mM NH4Cl so that Ph-L fusion was prevented, and that displayed many ferritin-labeled endosomes, revealed numerous examples of the fusion of electronlucent endosomes, revealed numerous examples of the fusion of electronlucent vesicles with phagosomes, whether containing M. microti bacilli or S. cerevisiae yeasts. Fusion was recognized by transfer of label and by morphological evidence of fusion in progress. The fusing vesicles were classed as endosomes, not NH4Cl-lysosomes, by their appearance and provenance, and because lysosome participation was excluded by the concurrent, NH4Cl-caused block of Ph-L fusion and associated lysosomal stasis. No evidence of such phagosome-endosome (Ph-E) fusion was observed in profiles from M phi treated with chloroquine, nor in those from normal, untreated M phi. NH4Cl-treated living M phi that had ingested yeasts at 37 degrees C, followed by endocytosis of lucifer yellow at 17 degrees C (to accumulate labeled endosomes and postpone label passing to lysosomes), were then restored to 37 degrees C. Fluorescence microscopy showed that as many as half of the yeast phagosomes (previously unlabeled) rapidly became colored. We inferred that this transfer was from endosomes (by Ph-E fusion) because Ph-L passage was blocked (by the NH4Cl). We conclude that NH4Cl induces Ph-E fusion at the same time as it suppressed Ph-L fusion. We discuss the mechanisms of these concurrent effects and suggest that they are independent; and we consider the implications of NH4Cl opening a direct route for endocytosed molecules to reach an intraphagosomal infection without involving lysosomes.

Ammonium Chloride↗
PubMed1991

Mycobacterial infection is an important infective complication in British Asian dialysis patients.

To define the extent and nature of mycobacterial infection in patients on an adult dialysis unit whose catchment population contains a large proportion of non-Caucasian subjects, a retrospective survey of all new patients accepted onto our maintenance dialysis programme between January 1987 and December 1989 was carried out. Twenty-six Asian, 13 Afro-Caribbean, two Oriental and 170 Caucasian patients were accepted onto the dialysis programme in the three-year recruitment period. Eight of the 26 Asian patients, but none of the others, had developed mycobacterial infection by the end of December 1990. One patient had a cerebral tuberculoma with miliary mottling on chest X-ray, one pulmonary tuberculosis, one tuberculous adenitis and 5 tuberculous peritonitis (four due to Mycobacterium tuberculosis and one Mycobacterium kansasii). All the patients had been living in the UK for an average of 15 (range 6-24) years, with no known recent exposure to tuberculosis. Five patients are now alive and well, one developed malabsorption following M. kansasii peritonitis, but two with tuberculous peritonitis died before treatment could be instituted. Mycobacterial infections were associated with a high level of mortality and morbidity. No Asian patient developed mycobacterial infection during post-transplant immunosuppressive therapy in the study period, probably because of the routine anti-tuberculous chemoprophylaxis employed in this group of patients. The diagnosis of mycobacterial infection should be suspected when an Asian dialysis patient develops a pyrexia of unknown origin. It is likely, though not proven, that anti-tuberculous chemoprophylaxis might reduce this high incidence of tuberculous infection in Asian dialysis patients.

Adult↗
PubMed1991

Disease pattern in the elderly.

Analysis of the disease pattern in elderly patients was carried out over a seven-year period (1980-1986) on the medical wards of the University College Hospital, Ibadan, Nigeria. A total of 2,676 patients were reviewed. Cardiac, neoplastic and infective diseases accounted for the most common diseases. Early diagnosis and treatment as well as improvement in preventive medicine are suggested ways to a vigorous healthy old age with concomitant reduction of morbidity and mortality in this age group.

Aged↗
PubMed1991

Diseases of the middle-aged population: a study of 4,184 patients.

Within a seven-year period (1980-1986), 4,184 Nigerians aged 31-60 years were admitted into the medical wards of the University College Hospital, Ibadan, Nigeria. The clinical notes of these patients were analysed and the principal diagnoses for each case was coded in accordance with the International Classification of Diseases. Cardiovascular, infective, neoplastic and neuro-muscular diseases constituted the commonest disorders being 70pc of all diseases at admission in this age group. The relevance of these findings in the light of the pattern of diseases elsewhere is discussed.

Age Factors↗
PubMed1990

Tubular lysosomes and their drug reactivity in cultured resident macrophages and in cell-free medium.

Lysosomes were assessed in normal living resident mouse peritoneal macrophages, using mainly phase-contrast microscopy (PCM), darkfield microscopy (DFM), and fluorescence microscopy (FM) after terminal acridine orange (AO) staining; these procedures avoided dyes during experimentation. After a few hours of culture a variable proportion of the normal spherical lysosomes began to assemble in a linear fashion. Fully formed tubular structures, with appearances generally recognized as characteristic of tubular lysosomes (TL), could be seen by PCM or, after labeling, by FM, at 2-5 days (best usually at 4-5 days). This peak was followed by a reduction, and at 8-10 days most of the TL had disappeared, leaving only spherical lysosomes. Renewal of the medium at this stage was followed by a temporary reappearance of TL, suggesting that the medium was a major factor in their initial development also. Formation of TL was enhanced by chloroquine (Cq), though to a lesser degree than by phorbol ester (PMA); in contrast NH4Cl (like Cq a weakly basic amine) caused their disassembly into spherical lysosomes. Manual disruption of the monolayer macrophages enabled TL to be transferred to a cell-free medium, in which they remained apparently stable for several hours. Two known microtubule depolymerizers caused disassembly of TL in the intact cells, reinforcing the idea that the TL are associated with the cytoplasmic microtubule (MT) system; but these agents were inactive in vitro, suggesting that disorganization of the system was responsible for this change.(ABSTRACT TRUNCATED AT 250 WORDS)

Ammonium Chloride↗
PubMed1990

Abnormal mucociliary action in asthma and bronchiectasis.

Nasal mucociliary clearance (NMCC) time was measured in four groups of patients: asthmatics with allergic rhinitis, asthmatics without rhinitis, bronchiectasis and normal subjects. The saccharin method was used for the study. The NMCC time was prolonged significantly in the asthmatic groups and group with bronchiectasis when compared with control subjects (P less than 0.001). It is likely that the impaired mucociliary clearance is due to a combination of mucus abnormality and ciliary malfunction.

Adult↗
PubMed1988

Polyanionic agents inhibit phagosome-lysosome fusion in cultured macrophages: a reply to the suggestion of Goren, Vatter, and Fiscus to the contrary.

The paper and review by Goren et al. (J. Leukocyte Biol. 41, 111, 1987) contain serious objections to the reports from several laboratories on the pattern of fusion of secondary lysosomes with phagosomes (yeasts being predominantly the target) in polyanion-treated macrophages; these reports had concluded that the polyanions were inhibitors of this fusion. The main objection by Goren et al. is to the alleged misuse of electron microscopic (EM) lysosome markers; many instances of phagosome-lysosome (P-L) fusion in the treated cells have therefore been missed. The central argument is that 1) the "hydrocolloid" properties of certain of these polyanions hinder the passage of the enmeshed marker from lysosome to phagosome after their fusion and 2) this hindrance is mistakenly interpreted as indicating that fusion has not taken place, thus giving rise to the belief that the polyanions can inhibit fusion. In reply, we explain that we score as P-L fusion any instance of marker (ferritin) being seen anywhere in a fused phagosome (phagolysosome). For this crucial reason, immobilisation of marker by a hydrocolloid polyanion, e.g., in lysosomal residue of phagolysosomes or just within phagosome membranes (as in Goren's Figs. 5 and 6 [8]), would not seriously threaten the marker distinction between fusion and nonfusion (with consequent underestimation of the former) and therefore would not invalidate the reports of a high incidence of nonfusion in polyanion-treated macrophages. Such inhibition of P-L fusion is supported by using as lysosomal label the nonpermeant fluorescent probe lucifer yellow (accepted by Goren et al. as a reliable indicator of fusion). Further support comes from the correlated inhibition of the saltatory movements of secondary lysosomes previously described; static lysosomes will have their contact with the phagosomes severely restricted. Another criticism is based on the failure of certain salient properties and functions of phagosomes to change significantly after polyanion treatment of the macrophages; these include intraphagosomal digestion, presence of lysosomal enzymes, and acidification. However, this indirect evidence can be accounted for alternatively by the operation of factors (primary lysosomes, endogenous acidification, etc.) not affected by the polyanionic block. We conclude that fusion inhibition by the polyanions is a real phenomenon, as previously reported, notwithstanding the hydrocolloid properties of some of them. Furthermore, an explanation based on hydrocolloid properties is questionable, since one or possibly two of the five main polyanionic agents appear not to be hydrocolloids.

Cells, Cultured↗
PubMed1987

Inhibition of phagosome-lysosome fusion in macrophages by certain mycobacteria can be explained by inhibition of lysosomal movements observed after phagocytosis.

We have investigated the mechanism of the inhibition of phagosome-lysosome (P-L) fusion in macrophages known to occur after infection by Mycobacterium tuberculosis and by the mouse pathogen Mycobacterium microti. We have used an M. microti infection and have studied, first, the saltatory movements of periphagosomal secondary lysosomes by means of visual phase-contrast microscopy (a similar use of the method having been previously supported by computer analyses). The movements became slow or static after ingestion of live but not of heat-killed M. microti. They were unaffected by a fusiogenic mycobacterium M. lepraemurium. Second, we studied the behavior of a normally fusiogenic unrelated organism, Saccharomyces cerevisiae, after its phagocytosis by cells already containing live M. microti ingested 18 h previously. We observed, using a fluorescent assay of fusion, that many of these yeast phagosomes now also failed to fuse with the lysosomes; in contrast, when the host M. microti had been heat killed the yeast phagosomes fused normally. These observations were extended by ultrastructural quantitative analyses of P-L fusion, which confirmed the nonfusion of phagosomes of live M. microti and, more particularly, the change to nonfusion from the normal fusion behavior of the separate phagosomes of accompanying yeasts. Third, we have assembled evidence against the likelihood that these M. microti-induced phenomena are nonspecific, i.e., secondary to a general depression of activity of heavily infected host cells. The evidence includes the feasibility of adjusting the degree of infection so as to facilitate visual assessment of organelle movements without the presence of detectable damage to the cells studied; the absence of lysosomal stasis after comparable infection with another mycobacterium of comparable virulence (M. lepraemurium); and the reversibility of the stasis. We conclude that inhibition of lysosome saltatory movements (and consequently its secondary effect on the associated yeasts) is a significant, specifically induced phenomenon. From these observations and considerations, therefore, in conjunction with the analogous inhibition of lysosomal movements in normal macrophages by some chemical inhibitors of P-L fusion, and our suggestion that this association is causally related, we now suggest that M. microti-induced focal lysosomal stasis is also the main means by which the inhibition of P-L fusion is brought about by this organism. This concept is strengthened by the observations on S. cerevisiae, which provide strong evidence that stasis can cause suppression of fusion.

Animals↗
PubMed1979

Effects of anionic inhibitors of phagosome-lysosome fusion in cultured macrophages when the ingested organism is Mycobacterium lepraemurium.

The mouse pathogen Mycobacterium lepraemurium is readily phagocytosed by cultured mouse peritoneal macrophages. Ingestion is normally followed by fusion between phagosomes and lysosomes. The influence of some aninonic compounds known to inhibit fusion in other systems was investigated by transmission electron microscopy after ingestion of M. lepraemurium. Fusion was markedly, although temporarily, inhibited by suramin and moderately inhibited by poly-D-glutamic acid. The effects are, however, not sufficient to permit these agents to be used to study the long-term effects of shutting off the secondary lysosome-phagosome fusion system in cultured macrophages infected with M. lepraemurium.

Animals↗
PubMed1977

BCG and vole bacillus vaccines in the prevention of tuberculosis in adolescence and early adult life.

The Medical Research Council's trial of BCG and vole bacillus vaccines in the prevention of tuberculosis in Great Britain has ended after 20 years' follow-up of the 54 239 participants, who were aged 14 to 15 years when the entered the trial in 1950-2. Participants who were tuberculin positive on entry were left unvaccinated; those who were tuberculin negative were allocated at random to an unvaccinated or to a vaccinated group. The protective efficacy of each of the two vaccines, among those initially tuberculin negative, was 84% during the first five years, and gradually decreased, averaging 77% for each vaccine over the whole period. The incidence of tuberculosis decreased substantially in all groups during the trial, however, and of the total of 610 cases of tuberculosis only 27 developed between 15 and 20 years. Thus we cannot make a reliable assessment of efficacy during this final period. The prevalence and incidence of tuberculosis in Great Britain have decreased radically since this trial began. The expected benefit from large-scale BCG-vaccination of children is now far less, and may decrease further if the incidence of tuberculosis continues to decline.

Adolescent↗