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T H Thomas

Publications and source records attributed to T H Thomas.

At least 37 records · Page 2Linked to original sources
PubMed1998

Lack of function of an N-ethylmaleimide-sensitive thiol protein in erythrocyte membrane of autosomal dominant polycystic kidney disease.

The polycystic kidney disease 1 (PKD1) gene product polycystin has been predicted to be an integral membrane protein involved in cell-cell and cell-matrix interactions. The erythrocyte membrane fluidity in autosomal dominant polycystic kidney disease (ADPKD) patients is increased, and this may be due to a membrane cytoskeletal abnormality. The abnormal erythrocyte sodium-lithium countertransport kinetics in-ADPKD are related to an altered thiol protein in the cytoskeleton. The possibility that a similar thiol protein abnormality causes the increased erythrocyte membrane fluidity in ADPKD was investigated. The membrane fluidity of intact erythrocytes from 12 ADPKD patients and 12 healthy control subjects was assessed from the fluorescence anisotropies of 1,6-diphenyl-1,3,5-hexatriene (DPH) and trimethylammonium-diphenyl-hexatriene (TMA-DPH). The effect on membrane fluidity of N-ethylmaleimide (NEM), cytochalasin D, heating at 48 degrees C for 20 min, or more specifically, liposomes containing antibodies to actin or ankyrin, was determined. In erythrocytes from healthy control subjects, the fluorescence anisotropy of DPH (mean +/- SEM: 0.223 +/- 0.001) was decreased after treatment with NEM (0.200 +/- 0.003, P < 0.001), cytochalasin D (0.206 +/- 0.006, P < 0.001), heating (0.199 +/- 0.002, P < 0.001), and antibodies to actin (0.194 +/- 0.002, P < 0.001) or ankyrin (0.196 +/- 0.002, P < 0.001). The TMA-DPH anisotropy (0.279 +/- 0.001) was also decreased after treatment with NEM (0.264 +/- 0.001, P < 0.001), cytochalasin D (0.264 +/- 0.001, P < 0.001), heating (0.265 +/- 0.001, P < 0.001), and antibodies to actin (0.262 +/- 0.002, P < 0.001) or ankyrin (0.262 +/- 0.002, P < 0.001). NEM had no additional effect on the other treatments, suggesting that its target thiol protein was associated with the cytoskeleton. In untreated erythrocytes from ADPKD patients, fluorescence anisotropies of both DPH and TMA-DPH were reduced, and none of the treatments altered the anisotropy of either DPH or TMA-DPH. In ADPKD, a cytoskeletal thiol protein is abnormal and possibly explains abnormal lipid bilayer properties and transport protein function in erythrocytes in this disease.

Actins↗
PubMed1997

Na-Li countertransport kinetics in the relatives of hypertensive patients with abnormal Na-Li countertransport activity.

Familial factors are believed to be important in determining the high sodium-lithium countertransport activity (defined as >0.40 mmol Li/(h x l cell) at external sodium concentration of 140 mmol/L (Nae 140)) which is observed in a proportion of patients with essential hypertension. However, environmental factors such as pregnancy and dyslipidemia also affect activity. High sodium-lithium countertransport activity (Nae 140) in essential hypertension is mainly due to a low Michaelis constant (Km) and is associated with a high Vmax/Km ratio. In contrast, dyslipidemias affect Vmax. This study aimed to determine if there was evidence that Km and Vmax/Km ratios are influenced by familial factors. Sodium-lithium countertransport kinetics were measured in the 47 first degree relatives of 12 hypertensive probands with abnormal sodium-lithium countertransport kinetics and 35 normotensive control subjects. Sodium-lithium countertransport was measured as Na-stimulated Li efflux from lithium loaded erythrocytes. The relatives had significantly reduced Km and increased Vmax/Km compared to normal subjects. Eleven relatives had high sodium-lithium countertransport activity (Nae 140), associated with low Km and high Vmax/Km. The 14 relatives that were hypertensive had abnormalities of sodium-lithium countertransport kinetics. The results of this study suggest that familial factors are important in determining the Km and Vmax/Km of sodium-lithium countertransport activity. Studies aimed at determining the inheritance of sodium-lithium countertransport and its use as an intermediate phenotype of essential hypertension must measure its kinetic determinants to reduce the risk of confounding effects from other variables.

Adult↗
PubMed1997

Thiol group modulation of sodium-lithium countertransport kinetics in diabetic nephropathy.

Abnormal erythrocyte sodium-lithium countertransport (Na-Li CT) activity, traditionally measured at a single sodium concentration of 140 mmol.l-1 (V140), may represent an inherited risk marker for diabetic nephropathy. The membrane defect underlying this association is poorly understood, though modulation by key protein thiol groups appears to be important in essential hypertension. To improve understanding of this abnormality, Na-Li CT kinetics in untreated erythrocytes and after thiol group alkylation with N-ethylmaleimide were investigated in 18 subjects with diabetic nephropathy, 20 normoalbuminuric insulin-dependent diabetic (IDDM) subjects and 18 non-diabetic individuals. Using the traditional assay, V140 was similar in subjects with diabetic nephropathy compared to IDDM control subjects vs 0.311 (0.152-0.475) (0.247 (0.111-0.498) mmol Li.h-1.l erythrocytes-1). Kinetic parameters were abnormal in subjects with diabetic nephropathy compared with diabetic and non-diabetic control subjects with both Vmax (maximal Na-Li CT activity) (0.454 (0.257-0.963) vs 0.338 (0.183-0.972) vs 0.332 (0.213-0.603) mmol Li.h-1.l erythrocytes-1, p < 0.05), and Vmax/Km(So) ratio, reflecting ion association (6.03 (2.3-9.6) vs 4.73 (2.0-10.4) vs 4.48 (1.5-7.1), p < 0.01), significantly higher. N-ethylmaleimide decreased K(m)(So) and Vmax abolishing differences in Vmax/Km(So) ratio between groups (2.45 (1.18-4.21) vs 2.23 (0.96-4.3) vs 2.44 (1.4-3.7), but enhancing the differences in Vmax (0.186 (0.090-0.315) vs 0.120 (0.051-0.256) vs 0.128 (0.080-0.206) mmol Li.h-1.l erythrocytes-1, p < 0.0001). Of subjects with diabetic nephropathy, 78% were outside the 75th percentile of the non-diabetic control subjects when Vmax and Vmax/Km(So) ratio were combined, compared to 20% of the normoalbuminuric control subjects. We conclude that the traditional assay, V140, is poor at detecting individuals with diabetic nephropathy. Study of the kinetic parameters of the transporter, including thiol group modulation, suggests that increased ion association, Vmax/Km(So) ratio may represent the inherited defect and improves identification of subjects with diabetic nephropathy.

Adult↗
PubMed1997

Insulin resistance in adult polycystic kidney disease.

Adult polycystic kidney disease (APKD) is a common hereditary disease with renal and extra-renal manifestations. There are at least three genes responsible for this disease. The polycystic kidney disease 1 (PKD1) gene product is a membrane protein involved in cell-cell and cell-matrix interactions and has a widespread tissue distribution. Abnormal membrane fluidity in erythrocytes from APKD patients is due to altered membrane proteins. Membrane fluidity of mononuclear cells is related to whole body insulin sensitivity. Insulin sensitivity might therefore be disturbed in APKD if the erythrocyte membrane abnormality is also present in other cells. Therefore, we investigated insulin sensitivity in 15 APKD patients and 20 normal subjects matched for age and sex. Insulin sensitivity was assessed by a short insulin tolerance test to derive the first-order rate constant for the disappearance of glucose (Kitt) and mononuclear leukocyte membrane fluidity was measured by fluorescence anisotropy. The Kitt value (% mmol.liter-1.min-1) was lower in APKD patients than in normal subjects [median (range) 2.2 (1.5 to 6.3) vs. 4.1 (2.0 to 5.4). P < 0.001]. Fasting plasma insulin concentrations were negatively correlated with the Kitt values (r = -0.66, P < 0.001). Core region anisotropy was significantly lower (higher fluidity) in leukocytes from APKD patients [mean (SEM) 0.164 (0.003) vs. 0.174 (0.001), P < 0.001]. Insulin sensitivity was positively correlated with the fluorescence anisotropy of the core region of leukocyte membranes (r = 0.81, P = 0.0001). In conclusion, APKD patients were insulin resistant and some patients were hyperinsulinemic, which may indicate increased cardiovascular risk. The cellular basis of the insulin resistance may be directly related to the proteins causing the disease or to the general change in membrane properties.

Adolescent↗
PubMed1996

Abnormal sodium-lithium countertransport kinetics in immunoglobulin A nephropathy patients and the families: association with hypertension.

Immunoglobulin A nephropathy (IgAN) is the most common primary glomerulonephritis worldwide. There is an increased prevalence of hypertension, which is an important risk factor for progressive renal impairment, in patients with IgAN. Changes in sodium-lithium countertransport (Na/Li CT) kinetics, particularly high Vmax/Km, have been shown in familial essential hypertension. In this study a high Vmax/Km was observed in IgA probands with hypertension and/or progressive renal impairment. Mean blood pressure was higher in the first-degree relatives of patients with IgAN compared with relatives of normotensive IgAN probands. These hypertensive relatives had an increased Vmax/Vm ratio and a low Km of Na/Li CT. There is a strong correlation of Vmax/Vm (r = 0.82) between the IgA probands and their first degree relatives, suggesting strong familial factors contributing to this Na/Li CT kinetic parameter. An increased Vmax/Km ratio of Na/Li CT seems to be a better marker for familial hypertension than Km alone and may be useful in identifying those patients who are at a greater risk of developing hypertension.

Adult↗
PubMed1996

Erythrocyte membrane fluidity in adult polycystic kidney disease: difference between intact cells and ghost membranes.

In adult polycystic kidney disease (APCKD) the cytoskeleton of renal tubular cells is abnormal. In erythrocytes the cytoskeleton affects the fluidity of membrane lipids. The authors determined fluorescence anisotropy in intact erythrocytes and erythrocyte ghosts in 12 APCKD patients and 12 normal subjects. In APCKD whole erythrocytes had a much lower core-region anisotropy, which indicated higher membrane fluidity than normal (mean 0 center dot 175 vs. 0 center dot 224, P < 0 center dot 01). This abnormality was not detected in erythrocyte ghosts, which suggests that preparation of ghosts altered membrane lipid organization. This could be directly due to ghosting or secondary to the loss of cytoskeletal effects, which may be abnormal in APCKD.

Adolescent↗
PubMed1996

Increased mononuclear cell membrane fluidity and increased B lymphocyte HLA class I expression in IgA nephropathy.

BACKGROUND: The functions of membrane bound proteins are regulated by the physical properties of the cell membrane. Lymphocyte dysfunction in IgA nephropathy may therefore be related to abnormal cell membrane fluidity. In this study we have examined peripheral blood mononuclear cell membrane fluidity and the expression of HLA antigens on the surface of lymphocytes in IgA nephropathy subjects compared to normal controls and disease controls. METHODS: Twenty-two IgA nephropathy subjects with normal or mildly elevated serum creatinine (serum creatinine RESULTS: Fluorescence anisotropy for diphenylhexatriene of mononuclear cells from IgA nephropathy subjects was significantly lower compared to mononuclear cells from normal control subjects and disease control subjects, indicating higher membrane fluidity (median values, IgA nephropathy 0.161; normal control 0.175; disease control 0.175; P<0.001 and P<0.001). Fluorescence anisotropy for trimethylammonium-DPH was lower in the IgA nephropathy groups compared to the normal control group (median values, IgA nephropathy 0.268; normal control 0.274; P<0.001), but not significantly different compared to the disease control group (0.272). HLA class I expression on the surface of B lymphocytes was significantly higher in the IgA nephropathy group compared to the normal control and disease control groups (median values, IgA nephropathy 30364, normal control 15495; disease control 16907; P=0.0001 and P=0.002 respectively). CONCLUSION: This study provides evidence of abnormal cell membrane architecture and increased HLA class I expression in Iga nephropathy.

Adult↗
PubMed1996

Erythrocyte ion and water balance and membrane potential in the puerperium of normal pregnancy.

OBJECTIVE: To examine the balance of erythrocyte ions and water during the rapid changes in plasma osmolality in the early puerperium, and during the subsequent period of sustained readjustment. DESIGN: A serial study from the third trimester of pregnancy to 20 weeks after delivery. PARTICIPANTS: Thirty-five primiparous women who had experienced no antenatal complications. MAIN OUTCOME MEASURES: Plasma osmolality, erythrocyte hydration, potassium, chloride and sodium were measured and nondiffusible ion content and erythrocyte membrane potential calculated. Plasma sodium, potassium and chloride were also measured. RESULTS: During the first week after delivery plasma osmolality increased (280 (SEM 0.52)-289 (SEM 0.64) mosmol/kg; P < 0.001) but erythrocyte hydration did not decrease (2.060 (SEM 0.018)-2.067 (SEM 0.021) 1/kg dry cells) because of an increase in total cell osmole content (577 (SEM 5.31)-597 (SEM 6.15) mosmol/kg dry cells; P = 0.001). This increase included nondiffusible anions, chloride and potassium. These changes in ionic balance did not affect membrane potential. After the first week of the puerperium and up to the 20th week, plasma osmolality was stable but erythrocyte osmole content and hydration both decreased. This was due to a decrease in nondiffusible anions and potassium with a smaller increase in chloride leading to a decrease in membrane potential (-14.31 (SEM 0.34)mV to -12.66 (SEM 0.28)mV; P < 0.001). CONCLUSIONS: A rapid increase in intracellular osmoles can occur in the mature erythrocyte and probably precedes the decrease in plasma osmolality in the puerperium. Changes in erythrocyte homeostasis in the first week of the puerperium can be accounted for by alterations in nondiffusible anions. After the first week of the puerperium it appears that the functional organisation of the membrane is changing.

Erythrocyte Membrane↗
PubMed1996

Thiol group control of sodium-lithium countertransport kinetics in uraemia: evidence of a membrane abnormality affected by haemodialysis.

Uraemia affects erythrocyte metabolism and membrane function but no consistent effect on Na/Li countertransport (CT) has been reported. We report only small differences in Na/Li CT at 150 mmol/l Na over haemodialysis, but major differences in other properties of Na/Li CT. The Km for external sodium and Vmax both increased during haemodialysis but the Vmax/Km ratio, which was greater than normal, was not affected. The thiol reagent, N-ethylmaleimide (NEM), which causes a decrease in Km and Vmax in normal subjects, had no effect on Km in the predialysis erythrocytes. After haemodialysis, the sensitivity of Na/Li CT to NEM was improved. The changes in Na/Li CT kinetics were not related to changes in membrane lipid fluidity or plasma lipids. These observations suggest that uraemia affects a thiol group that controls Na/Li CT kinetics and that haemodialysis temporarily improves this aspect of membrane function.

Aged↗
PubMed1996

Increased susceptibility to membrane lipid peroxidation in renal failure.

The susceptibility of membrane lipids to peroxidation in renal failure was investigated. A fluorescent polyunsaturated fatty acid probe, cis-parinaric acid, showed that mononuclear leucocytes from both chronic renal failure and haemodialysis patients had increased susceptibility to membrane lipid peroxidation compared with normal controls. The effect of oxidant stress on the physical properties of the cell membrane was also investigated by measuring fluidity with the probes diphenylhexatriene and trimethylaminodiphenylhexatriene. Oxidant stress caused a significant increase in membrane fluidity of mononuclear leucocytes from normal controls, which was greater than that seen in renal failure. The effect of oxidant stress on the physical properties of the cell membrane was not simply determined by the susceptibility of the membrane lipids to peroxidation.

Cell Membrane↗
PubMed1995

Modification of erythrocyte Na+/Li+ countertransport kinetics by two types of thiol group.

Erythrocyte Na+/Li+ countertransport activity is decreased by reagents that react with thiol groups. An understanding of the role of these groups in control of Na+/Li+ countertransport may help to explain its association with disease states. The effect of thiol reactive agents on the kinetic parameters of Na+/Li+ countertransport has not previously been described. In choline medium, N-ethylmaleimide (NEM) and iodoacetamide (IAamide) cause a rapid decrease of about 40% in Km for external sodium (Km(So)) that is complete in 10 s with a much smaller change in Vmax and an increase in the Vmax/Km ratio. In Na medium, NEM and IAamide both cause a rapid decrease in Km(So) and Vmax. With NEM the partial reduction in Vmax is complete in 100s although the NEM is sufficient to reduce Vmax up to 15 min. With IAamide the decrease in Vmax is initially slower but it continues apparently towards complete inhibition. These results indicate at least two types of thiol group controlling Na+/Li+ countertransport kinetics. The type 1 thiol reaction is Na independent and causes an increase in the apparent rate constant for Na association with the unloaded carrier so that Vmax/Km rises and Km(So) decreases. The type 2 thiol reaction is facilitated by Na at the outside ion-binding site and causes a decrease in Vmax, possibly by total blockage of carriers with IAamide but by a different mechanism with NEM such as reduced turnover rate.

Antiporters↗
PubMed1995

Abnormal effect of thiol groups on erythrocyte Na/Li countertransport kinetics in adult polycystic kidney disease.

BACKGROUND: Evidence suggests that in adult polycystic kidney disease (APCKD) there is abnormal membrane organisation that involves cytoskeletal proteins and affects ion-transport proteins. The possibility of detecting a membrane organisation defect in erythrocytes from APCKD patients was investigated. METHODS: Na/Li countertransport (CT) kinetics were measured in erythrocytes from APCKD patients compared with normal controls. The modulation of Na/Li CT kinetics by key membrane thiol groups and by dissociation of spectrin by heating was studied. RESULTS: In erythrocytes from APCKD compared to normal subjects the Km of Na/Li CT was lower (64 SEM 3v 90 SEM 3 mmol Na/l, P < 0.001) and Vmax/Km was higher (6.8 SEM 0.6v 5.3 SEM 0.4 (x 19(-3)), P < 0.05). In erythrocytes from normal subjects, after N-ethylmaleimide (NEM) treatment in choline medium, Km of Na/Li CT was reduced and Vmax/Km increased (59 SEM 5, 8.2 SEM 0.4 (x 10(-3)), P < 0.001) but there was no effect on these kinetic parameters in erythrocytes from APCKD. The effect of heating was similar to that of NEM in choline medium. In normal controls after NEM treatment in sodium medium both Km and Vmax were reduced whereas in APCKD Vmax was reduced but Km was unchanged. CONCLUSIONS: In APCKD the effect of a key membrane thiol group on Na association with Na/Li CT was absent and the effect of spectrin dissociation was similarily abnormal. A second thiol group effect on the Vmax of Na/Li CT was normal. The results are consistent with a thiol-protein linked to the spectrin cytoskeleton that modulates Na/Li CT and is abnormal in APCKD.

Adolescent↗
PubMed1995

Sulphydryl group control of sodium-lithium countertransport kinetics: a membrane protein control abnormality in essential hypertension.

Erythrocyte sodium-lithium countertransport (SLC) is an obligatorily coupled equimolar exchange of intracellular sodium or lithium with extracellular sodium or lithium. SLC is partially inhibited by N-ethylmaleimide (NEM) but only when a transported ion (sodium of lithium) is present in the extracellular medium. In essential hypertensive patients with a strong family history of hypertension the Km of SLC for extracellular sodium was lower and Vmax tended to be higher than in normal controls, but the ratio Vmax/Km gave a much clearer distinction between the two groups. After NEM treatment, the remaining SLC activity in normal individuals had a lower Vmax and Km for sodium but Vmax/Km was not affected. In essential hypertensives the remaining SLC activity after NEM again had lowered Vmax and Km but in these patients the Vmax/Km was much lower than in untreated erythrocytes and was then the same as in normal controls. On the assumption that NEM reacts with a -SH group on a membrane protein that regulates SLC, and that the ratio Vmax/Km reflects a rate constant for binding extracellular sodium to the unloaded carrier, the results suggest that (a) essential hypertensives have an increased rate of sodium binding to the transporter and (b) this is due to abnormal behaviour of a membrane -SH group.

Adolescent↗
PubMed1995

Sodium-lithium countertransport kinetics in IgA nephropathy: relation to plasma lipids and renal impairment.

We have investigated the activity and kinetics of sodium-lithium countertransport (SLC) in patients with IgA nephropathy and their relationship to plasma lipids. Standard SLC activity, the Michaelis constant (Km) and maximum velocity (Vmax) were measured in patients who had IgA nephropathy with either normal serum creatinine (IgA-NRF), or raised serum creatinine (IgA-IRF), and normal subjects (NC). The standard SLC activity was raised in hypertensive patients with IgA-NRF due to a raised Vmax in association with hyperlipidaemia. The Km was significantly lower and Vmax also tended to be lower in IgA-IRF. Km and Vmax were not different in IgA-NRF compared with the NC. There was no difference in the mean standard SLC activity between all three groups. The low Km and low Vmax resulted in a normal standard SLC activity being observed in IgA-IRF which is similar to the situation we have observed in a proportion of diabetic patients with nephropathy. The low Km in patients with IgA nephropathy may be due to inheritance associated with familial essential hypertension or to an acquired change of the kinetics related to a change in the environment of the plasma membrane during the development of renal impairment.

Adult↗
PubMed1994

Erythrocyte sodium-lithium countertransport activity is related to membrane fluidity in IDDM patients.

Sodium-lithium countertransport (SLC) activity at a standard physiological sodium concentration is raised in uncomplicated IDDM, for which the kinetic mechanism is a raised maximum velocity (Vmax). Diabetic patients with nephropathy do not have raised values for Vmax but a low Michaelis constant (km). Transporter activity could be influenced by its membrane lipid environment. This was assessed in 21 control subjects, 32 uncomplicated diabetic patients, 17 patients with diabetic nephropathy and 11 patients with non-diabetic nephropathy by measuring the fluorescence anisotropy of DPH and TMA-DPH to assess different membrane regions. Standard SLC was higher in all the patient groups compared to the control subjects: 0.307 +/- 0.020 mmol Li/h x 1 cells in uncomplicated IDDM; 0.300 +/- 0.032 in diabetic nephropathy patients and 0.276 +/- 0.019 in non-diabetic nephropathy patients vs 0.216 +/- 0.011 mmol Li/h x 1 cells in control subjects (p < 0.001, p < 0.05, p < 0.05, respectively). This was due to raised Vmax values in the uncomplicated group: 0.528 +/- 0.035 vs 0.385 +/- 0.022 mmol Li/h x 1 cells in control subjects (p = 0.001) and low values for km in the diabetic nephropathy group: 58 (27-170) vs 106 (81-161) mmol/l in control subjects (p < 0.001). Raised SLC in the non-diabetic nephropathy group was largely due to raised Vmax: 0.460 +/- 0.030 mmol Li/h x 1 cells; p = 0.053, with no difference in km: 99.5 (74-137).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1994

A serial study of erythrocyte sodium pump kinetics and sodium content in the puerperium.

OBJECTIVE: Our purpose was to describe the alterations in erythrocyte sodium pump kinetics and sodium content occurring during the puerperium. STUDY DESIGN: Twelve healthy primigravid women were studied serially from late pregnancy until 20 weeks after delivery. Erythrocyte sodium pump rate constant, maximum velocity, and sodium affinity were calculated from the ouabain-sensitive sodium flux measured in whole blood and in erythrocytes in which sodium content had been altered with the ionophore nystatin. The Student t test was used to compare the regression coefficients of the values plotted against log time for specific periods. RESULTS: The sodium pump rate constant, maximum velocity, and sodium affinity were lower 20 weeks after delivery than in late pregnancy (0.339 +/- 0.018 vs 0.399 +/- 0.016/hr, 7.02 +/- 0.08 vs 9.98 +/- 0.078 mmol/kg/hr, 2.65 +/- 0.21 vs 3.16 +/- 0.20 mmol/kg). The decrease in the rate constant commenced after 4 days of the puerperium, whereas the decrease in maximum velocity and Michaelis-Menten constant did not commence until after 2 weeks. Erythrocyte sodium content was greater 20 weeks after delivery than in late pregnancy (4.71 +/- 0.20 vs 4.14 +/- 0.15 mmol/kg cells) and the increase was gradual over the time studied. CONCLUSIONS: After delivery the rate constant of the sodium pump measured in plasma and the erythrocyte sodium content changed before any significant alteration in the maximum velocity of the pump. The return of sodium pump function to the nonpregnant state continues beyond 6 weeks after delivery.

Erythrocytes↗