Biomedical subjects
J Lowe
Publications and source records attributed to J Lowe.
Osmotic activation of a Na(+)-dependent Cl-/HCO3- exchanger.
In many systems, osmotically induced cell shrinkage activates the Na+/H+ exchanger. To assess the role of H(+)-extruding transporters in the response to osmotic shrinkage in vascular smooth muscle (VSM) and Chinese hamster ovary (CHO) cells, intracellular pH (pHi) was measured with 2',7'-bis(carboxy-ethyl)-5(6)- carboxyfluorescein-acetoxymethyl ester (BCECF-AM) after exposing cells to hypertonic medium. In nominally HCO(3-)-free medium, addition of 200 mM sucrose caused pHi to increase 0.33 pH unit on average in VSM cells but only 0.13 pH unit in CHO cells. Permeant solutes failed to increase pHi significantly. Cytochalasin B (1-20 microM), colchicine (1-10 microM), Ca2+ removal, and downregulation of protein kinase C activity did not affect osmotic activation of H+ extrusion in either cell type. Additional work was carried out to determine why osmotic activation of H+ extrusion was less in CHO than in VSM cells. In CHO cells, the osmotically induced delta pHi was only weakly sensitive to amiloride, suggesting that osmotic forces may activate an H+ transport system other than Na+/H+ exchange. In the presence of 10 mM HCO3-, osmotically induced delta pHi decreased by 60% in VSM cells but increased by 50% in CHO cells compared with the delta pHi in HCO(3-)-free medium. Lastly, removal of extracellular Cl- did not affect osmotically induced delta pHi in VSM cells but completely abolished the response in CHO cells. We conclude that in VSM cells osmotically induced changes in pHi are mediated by Na+/H+ exchange, whereas in CHO cells they are most likely mediated by a Na(+)-dependent Cl-/HCO3- exchanger.
Skeletal muscle lymphocytic vasculitis in systemic lupus erythematosus: relation to disease activity.
Lymphocytic vasculitis (LV) characterises systemic lupus erythematosus (SLE) and this potentially reversible lesion, which may be subclinical, may imply overt systemic disease activity. Needle quadriceps muscle biopsy was performed in 26 unselected patients with SLE and the presence of LV in these muscle specimens was compared with SLE disease activity scored using the British Isles Lupus Assessment Group Index (BILAG). Ten of the 22 patients with active disease showed evidence of LV compared with none of the four patients with inactive disease. In the patient group with LV, significantly higher ESR and urine neopterin values were found with P = 0.002 and P = 0.02, respectively compared with patients without LV. Features of vasculitis (as defined by BILAG) were also significantly more common in these patients (P = 0.005). None of the other parameters, including creatine kinase, were significantly different between the two patient subgroups. Thus, LV in needle quadriceps muscle biopsy specimens is a further valuable marker of disease activity in patients with SLE and might provide histological evidence of a systemic vasculitic process in a group of patients with diverse clinical manifestations.
Chicken B cells undergo discrete developmental changes in surface carbohydrate structure that appear to play a role in directing lymphocyte migration during embryogenesis.
The migration of progenitor cells to specific microenvironments is essential for the development of complex organisms. Avian species possess a unique primary lymphoid organ, the bursa of Fabricius, that plays a central role in the development of B cells. B cell progenitors, however, arise outside the bursa of Fabricius and, during embryonic development, must migrate through the vasculature to the bursa of Fabricius. In this report, we demonstrate that these progenitor B cells express the sialyl Lewis x carbohydrate structure previously shown to be a ligand for the selectin family of vascular adhesion receptors. Soon after migration to the bursa of Fabricius, B cell progenitors are induced to undergo a developmental switch and terminate the expression of sialyl Lewis x in a temporal pattern that correlates with the developmental decline in the ability of these cells to home to the bursa of Fabricius upon transplantation. The induction of the developmental switch in the glycosylation pattern of developing B cells requires the bursal environment. In addition, sialyl Lewis x carbohydrate determinants or structurally similar determinants on the surface of immortalized bursal lymphoid stem cells participate in the adherence of these cells to the vascular regions of the bursal microenvironment. These data demonstrate that the carbohydrate structure sialyl Lewis x is developmentally regulated during chicken B cell development and may facilitate the migration of B cell progenitors to the bursal microenvironment by serving as a ligand for a lectin-like adhesion receptor.
Follicular fluid levels of midazolam, fentanyl, and alfentanil during transvaginal oocyte retrieval.
OBJECTIVE: To investigate the time course of changes in follicular fluid (FF) concentrations of midazolam (Roche Products Ltd., Welwyn Garden City, United Kingdom), fentanyl (Janssen Pharmaceuticals Ltd., Wantage, United Kingdom), and alfentanil (Janssen Pharmaceuticals Ltd.) during ultrasound-guided transvaginal oocyte collection. STUDY DESIGN: Forty-five patients with tubal infertility were randomized to receive a bolus IV dose of midazolam, fentanyl, or alfentanil for sedation during ultrasound-guided transvaginal oocyte collection. Paracervical block with lignocaine was given for analgesia. Simultaneous blood and FF samples were drawn at 5-minute intervals after the bolus dose for analysis of drug levels. RESULTS: Data were obtained on 15 women receiving midazolam and fentanyl and on 13 women receiving alfentanil. Plasma levels of all agents rose to a peak and then fell in an exponential fashion as was expected. The FF levels of the agents continued to rise significantly to 25 minutes after the bolus dose, although the absolute level was low when compared with the blood level. There were no significant differences in fertilization or pregnancy rates in the three groups, but patient numbers were small. CONCLUSION: We conclude that midazolam, fentanyl, and alfentanil are found in FF after a single IV dose, but further investigation needs to be undertaken to investigate any potential influence on fertilization and implantation rates.
Human monoclonal anti-D secreting heterohybridomas from peripheral B lymphocytes expanded in the CD40 system.
Peripheral human B lymphocytes isolated from four immunised anti-D donors have been cultured in the CD40 system (Banchereau et al., 1991), prior to fusion to murine X63Ag8.653 plasmacytoma cells. High fusion efficiency was noted in all cases and immunoglobulin (IgG and IgM) was detected in nearly all heterohybrid containing wells. Only fusions using boosted donor lymphocytes generated specific anti-D secreting hybrids. A short period of culture of committed B cell precursors in the CD40 system before fusion appears to be both adequate and efficient in permitting generation of specific antibody-secreting hybrids.
A novel bioactivity assay for monoclonal antibodies directed against IgE.
A novel bioactivity assay has been developed to quantitate the biological activity of a humanized, monoclonal anti-IgE antibody (rhuMAbE25) in human whole blood. Heparinized blood specimens from prescreened healthy donors were sensitized for 2 h with a constant amount of human plasma containing IgE specific for ragweed and then challenged with ragweed allergen. Histamine was released in a dose-dependent fashion and reached plateau levels after 30 min. As expected, the release of histamine by ragweed allergen was time, temperature and Ca2+ dependent, and could be enhanced by the presence of 33% deuterium oxide. Allergen-triggered release could be inhibited by rhuMAbE25 with an effective dose range from 0.1 to 1 microgram/ml. Preincubation with other humanized MAbs, which exhibit 95% homology to rhuMAbE25 but differ in epitope specificity, failed to inhibit the ragweed-induced histamine release. Overall, this bioactivity assay has a low interassay variability (%CV) of 17% (n = 23) and can be readily modified to determine if rhuMAbE25 or other anti-allergy therapeutics are capable of blocking histamine release elicited by other allergens. Moreover, the assay can be used to confirm IgE-mediated allergic responses and to provide early information regarding safety and potential efficacy of therapeutics aimed at blocking IgE dependent responses.
Alpha-B crystallin in the normal human myocardium and cardiac conducting system.
The alpha crystallins are major protein components of the ocular lens and show both structural and functional homology with the family of small heat shock proteins. alpha B crystallin is also present in various extra-lenticular tissues, with a high concentration in cardiac muscle. In this study, the myocardium and conducting system from 15 adult and 25 fetal and infant hearts were examined by immunohistochemistry using a previously characterized antiserum to alpha B crystallin. Contractile myocardium showed moderate staining, with particular localization to Z bands and intercalated discs. Fibres of the sino-atrial and atrio-ventricular nodes and His bundle showed less intense staining than contractile fibres, whereas fibres of the left and right bundle branches showed more intense staining. This distribution is similar to that previously demonstrated for the intermediate filament desmin. This observation, together with currently available evidence, suggests that cardiac alpha B crystallin may play a role in protecting the cytoskeleton during cell stress. For practical purposes, immunostaining with alpha B crystallin greatly facilitates the identification of cardiac conducting fibres.
The expression of alpha B-crystallin in epithelial tumours: a useful tumour marker?
Alpha B-crystallin is a lens protein showing homology with small heat-shock proteins. We have previously demonstrated its expression in non-lenticular normal and diseased human tissues by immunostaining with a polyclonal antibody. In view of the expression seen in normal renal tubular epithelium, we have assessed the immunoreactivity of a variety of epithelial tumours, to determine the usefulness of alpha B-crystallin as a specific renal tumour marker. Carcinomas arising from tissues which normally express alpha B-crystallin, such as colo-rectal and thyroid carcinomas, showed a varying pattern and degree of immunoreactivity. The most consistently positive tumours, however, with typically strong cytoplasmic and cell membrane staining, were renal cell carcinomas, 90 per cent of which showed positive immunoreactivity. This pattern of staining, while not absolutely specific, is a useful aid to the diagnosis of renal carcinoma. When a metastic deposit or a small biopsy is being assessed, anti-alpha B-crystallin may be included in a panel of antibodies, the pattern of staining of which may direct the search for the primary site of the tumour.
New pathological findings in amyotrophic lateral sclerosis.
There have been recent developments in the pathology of sporadic ALS. A new filamentous neuronal inclusion body in ALS detected by immunohistochemical localisation of the protein ubiquitin has been characterised at the light microscopic and ultrastructural level and appears specific for the disease. The molecular composition of underlying filaments remains unresolved but the quest for this is a major aim in ALS research. Despite being a progressive degenerative process which primarily affects motor systems, ALS is now recognised to involve several non-motor systems and in long survivors affects many subcortical structures. There is also accumulating evidence that the neurodegenerative process underlying ALS may present as a non-motor clinical syndrome, particularly as a frontal lobe dementia with characteristic inclusions present in the non-motor cortex. Considering ALS as a multisystem disease rather than simply a disease of motor neurones has major implications for research into pathogenesis.
Endosome-lysosomes and neurodegeneration.
A number of the major human and animal neurodegenerative diseases, such as Alzheimer's disease and sheep scrapie, are characterised by deposits of amyloid, arising through incomplete breakdown of membrane proteins. Although our knowledge concerning these diseases is increasing, they remain largely untreatable. Recently, attention has focussed on the mechanisms of production of different types of amyloid and the likely involvement within cells of acid compartments called endosome-lysosomes. These organelles may be 'bioreactor' sites for the unfolding and partial degradation of membrane proteins to generate the amyloid materials. These subsequently become expelled from the cell, or are released from dead cells, and accumulate as pathological entities. Common features of the disease processes give new direction to therapeutic intervention.
Is there a common intracellular bioreactor in which amyloid formation is initiated in neurodegenerative diseases?
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Composite particle production in relativistic Au+Pt, Si+Pt, and p+Pt collisions.
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Muscle biopsy abnormalities in systemic lupus erythematosus: correlation with clinical and laboratory parameters.
OBJECTIVES: To investigate the incidence and significance of Type II fibre atrophy, vessel wall thickening, lymphocytic vasculitis and myositis in needle quadriceps muscle biopsies from patients with systemic lupus erythematosus (SLE) and their correlations with clinical and laboratory parameters. METHODS: Needle quadriceps muscle biopsies from 55 patients with SLE and 26 controls were prospectively examined. Clinical and laboratory parameters recorded at the time of muscle biopsy included arthralgia, arthritis, myalgia, proximal weakness, vasculitic rashes, Schirmer test, ENA antibodies, ESR, serum creatine kinase (CK) and plasma C3 degradation products. RESULTS: Abnormal muscle biopsies were significantly more frequent in patients with SLE compared with controls (P < 0.005). None of the controls had lymphocytic vasculitis and/or myositis. The difference in incidence between patients with SLE and controls for lymphocytic vasculitis was significant at P < 0.005. Due to the small number of SLE patients with myositis, the difference in incidence for this abnormal finding reached only P = 0.09. In the SLE patient group, lymphocytic vasculitis was associated with significantly higher ESR values (P = 0.007) and higher incidence of arthritis (P = 0.01); and appears to characterise a subset of patients with positive Schirmer tests, anti-Ro and/or anti-La antibodies. Raised serum CK was found to correspond with underlying myositis in patients with SLE and these patients also had an increased incidence of symptoms of proximal weakness and/or anti-RNP antibodies. In contrast, both Type II fibre atrophy and vessel wall thickening failed to correlate with any of the clinical and laboratory parameters studied and appear to be non-specific findings. CONCLUSIONS: Abnormal muscle biopsies are common in patients with SLE and the presence of lymphocytic vasculitis and/or myositis signify pathology in these patients. Histopathological abnormalities in needle quadriceps muscle biopsies are further valuable parameters in the assessment of patients with SLE.
An electron microscopic study of muscle capillary wall thickening in systemic lupus erythematosus.
Capillary vessel wall thickening may contribute to the muscle fibre atrophy and impaired pulmonary gas transfer notable in SLE/MCTD. Using electron microscopy we measured the thickness of unilaminar capillary basement membranes (CBM) and quantified basement membrane laminae and pericyte layers in the skeletal muscle biopsies of 31 patients with SLE, including 14 with MCTD, and 11 controls. Capillary basement membrane thickness was significantly increased in SLE compared with controls (P < 0.01) and was significantly associated with the level of C3dg (P < 0.01). Patients with MCTD had significantly greater layering of pericytes and laminae compared with patients with SLE alone (P < 0.0001 for both variables). Multiple stepwise regression analysis indicated that ANA titre was a good predictor of pericyte layering (R = 0.63, P = 0.003) and of basement membrane lamination (P < 0.0001), with steroid therapy having a significant negative association with the number of laminae (P < 0.02). A negative association between KCO and the number of layers of basement membrane (R = 0.51, P = 0.01) was documented. However, multiple regression analysis revealed no association between parameters of thickened vessel walls and the presence of muscle fibre atrophy.
Multidrug resistance gene (MDR 1) expression in neuro-axial tumours of children and young adults.
Drug resistance in many cancers outside the CNS has been associated with over-expression of the multidrug resistance gene (MDR1), which codes for the transmembrane efflux pump P-glycoprotein (Pgp). To determine whether tumours of the neuroaxis over-express MDR1 and to identify the site of Pgp expression we examined 50 tumour specimens from 46 children and young adults using immunocytochemistry. Pgp was not expressed by any neoplastic cells, but was detected in the endothelium of tumour blood vessels in 35 of the 50 samples (70%). 11/35 (31%) were Pgp positive in the majority of vessels, 11/35 (31%) in a proportion, but < 50% of vessels, and 13/35 (37%) in one or two vessels. Pgp was also detected in surrounding normal brain capillaries. MDR1 may play a role in the chemoresistance of neuro-axial tumours either by its expression in the normal blood-brain barrier or by forming a blood-tumour barrier. The proportion of vessels expressing Pgp may determine the degree of resistance.
Aortic valve replacement in adults after balloon aortic valvuloplasty.
BACKGROUND: Percutaneous balloon aortic valvuloplasty is limited by a high risk of procedural morbidity, transient clinical benefit, and a high restenosis rate. The management of patients with symptomatic aortic valve restenosis after percutaneous balloon aortic valvuloplasty is unclear. We hypothesized that aortic valve replacement would produce superior midterm survival compared with repeat balloon aortic valvuloplasty or medication alone in patients with symptomatic aortic valve restenosis after prior balloon aortic valvuloplasty. METHODS AND RESULTS: Baseline clinical, echocardiographic, and hemodynamic data were collected on 165 patients who underwent percutaneous balloon aortic valvuloplasty as treatment for symptomatic degenerative calcific aortic stenosis. In 144 of these patients (87%), aortic valve replacement was originally considered to carry excessive risk. The survival of three subgroups was calculated during a median follow-up period of 3.9 years (range, 1 to 6 years). Ninety-four patients (57%) had no further mechanical intervention (subgroup 1-BAV), 31 patients (19%) developed symptomatic aortic valve restenosis and underwent a repeat balloon aortic valvuloplasty (subgroup 2-BAV), and 40 patients (24%) subsequently underwent aortic valve replacement (subgroup BAV+AVR). Follow-up was 99% complete. Patients in subgroup BAV+AVR tended to be younger and have a lower prevalence of coronary artery disease or mitral regurgitation. Only 1 patient (2.5%) suffered a perioperative death during aortic valve replacement. The probability of survival 3 years from the date of the last mechanical intervention was 13% for subgroup 1-BAV, 20% for subgroup 2-BAV, and 75% for subgroup BAV+AVR. At the conclusion of follow-up, only 2 patients had symptoms of congestive heart failure or angina after aortic valve replacement. CONCLUSIONS: Aortic valve replacement may be performed with a low mortality rate, excellent palliation of symptoms, and prolongation of survival in selected high-risk patients with a history of previous balloon aortic valvuloplasty.
Diabetic microalbuminuria and cilazapril.
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