Search PubMed⌕ Search

Biomedical subjects

L L Ng

Publications and source records attributed to L L Ng.

At least 55 records · Page 3Linked to original sources
PubMed1997

Psychogeriatrics in South-East Asia.

A common phenomenon in South-East Asia is ageing of the population. This article describes the various stages of development of psychogeriatrics in Hong Kong, Singapore, Malaysia, Thailand, Indonesia and the Philippines. It is only in the last few years that more systematic development of psychogeriatric services has begun under the pressure of an ageing population. The model of service delivery in Hong Kong can serve as an example of development of psychogeriatric services in South-East Asia.

Aged↗
PubMed1997

Calcium-induced activation of the rat vascular myocyte Na+/H+ exchanger isoform-1.

An established intermediate phenotype of human hypertension and diabetic nephropathy is an elevation of Na+/H+ exchanger (NHE) activity, but the mechanism for this is unclear. This phenotype is maintained in vascular myocytes from the spontaneously hypertensive rat (SHR) compared with the normotensive Wistar Kyoto rat (WKY). Since intracellular calcium levels ([Ca2+]i) following agonist stimulation were elevated in cells from both hypertensive humans and SHR, we have examined the role of calcium-calmodulin (CaM) in the mechanism of increased NHE activity in vascular myocytes of SHR by determining the activity and phosphorylation state of NHE isoform-1 (NHE-1) in cells from SHR and WKY when [Ca2+]i was elevated by the ionophores A23187 or ionomycin. NHE activity was measured using fluorometry and NHE-1 phosphorylation by immunoprecipitating the exchanger from 32P-orthophosphate-labeled cells with a polyclonal NHE-1-specific antibody. The ionophore A23187 increased [Ca2+]i in both cell types to approximately 700 to 800 nmol x L(-1), and led to stimulation of NHE-1 activity only in WKY myocytes, with no effect on SHR cells. An inhibitor of CaM kinase II (KN-62) failed to abolish stimulation of NHE-1 by A23187 in WKY cells, and had no effect on unstimulated NHE-1 activity in both cell types. Ionomycin also elevated [Ca2+]i in both cell types to approximately 1,000 nmol x L(-1) and activated NHE-1 activity in only WKY cells. Activation of NHE-1 in WKY cells by an increased [Ca2+]i was not mediated by an increase in NHE-1 phosphorylation, whether in the presence or absence of KN-62. The elevated NHE-1 phosphorylation in SHR cells was not affected by elevated [Ca2+]i or KN-62. Calmodulin-agarose beads bound NHE-1 extracted from SHR cells to a lesser extent than that from WKY cells. We conclude that calcium-induced NHE-1 activation in WKY cells was not mediated by CaM kinase II. The elevated NHE-1 activity and phosphorylation of SHR cells was not further modulated by increased [Ca2+]i, and was also independent of CaM kinase II. Non-phosphorylation-dependent mechanisms of activation of NHE-1 may therefore be responsible for alterations of NHE-1 activity in these cells, such as the direct binding of CaM to NHE-1. This direct binding of CaM to NHE-1 may be impaired in SHR compared with WKY cells.

Animals↗
PubMed1997

Enhanced mitogen-activated protein kinase activity and phosphorylation of the Na+/H+ exchanger isoform-1 of human lymphoblasts in hypertension.

Increased activity of the Na+/H+ exchanger isoform-1 (NHE-1) is recognized as an intermediate phenotype for hypertension, but the basis for this association is unclear. We have previously demonstrated an increased phosphorylation of NHE-1 in lymphoblasts from hypertensives that was associated with increased cell proliferation. Due to the central importance of mitogen-activated protein kinases (MAPKs) in signaling cascades transducing responses from extracellular growth factors and hormones, we examined the activity of this kinase in a specific peptide phosphorylation assay. Cells from hypertensives showed a significant twofold enhancement of MAPK activity (P < .001). This was not associated with any increase in p42mapk and p44mapk protein content. There was no significant increase in the level of tyrosine phosphorylation of MAPK in cells from hypertensives. MAPK activity was correlated with NHE-1 activity (r(s) = .55, P < .01) and phosphorylation (r(s) = .51, P < .02). These findings suggest that the increased cell proliferation rate, NHE-1 activity, and phosphorylation of lymphoblasts from hypertensives may be associated with enhanced MAPK activity, suggesting upregulation of this signaling pathway in hypertension.

Adult↗
PubMed1997

Angiotensin II responses of vascular smooth muscle cells from hypertensive rats: enhancement at the level of p42 and p44 mitogen activated protein kinase.

1. Stimulation of the AT1 receptor by angiotensin II (AII) gives a larger mitogenic response in vascular smooth muscle cells from spontaneously hypertensive rats (SHR) compared to those from normotensive (WKY) controls. Here we investigated whether the p42 and p44 mitogen activated protein kinase (MAPK) pathway is differentially regulated in these cells by AT1 receptors. 2. We showed that there is a similar level of p42 and p44 MAPK immunoreactivity in the SHR and WKY derived cells. 3. However, by use of an antiserum specific for the tyrosine phosphorylated form of MAPK, and an assay with a nonapeptide MAPK substrate, we showed that AII (100 nM)-stimulated phosphorylation and activation of p42mapk and p44mapk are enhanced in the SHR derived cells. 4. This increased MAPK activity in SHR derived cells was also seen on protein kinase C activation with 100 nM phorbol myristate acetate (PMA). The size and time course of the response to PMA was the same as that to AII in each cell type. 5. The protein kinase C inhibitor Ro 31-8220 attenuated the early (2 min) phase of AII stimulation of MAPK activity and the entire stimulation caused by PMA. At longer times of AII stimulation both p42mapk and p44mapk were activated by an Ro 31-8220-insensitive mechanism. 6. Agonist or PMA stimulation of MAPK activity was inhibited by the tyrosine kinase inhibitor genistein. AII stimulated tyrosine protein phosphorylation to a greater degree in SHR than WKY cells. 7. These results show that the MAPK response of SHR derived cells is increased over that of WKY cells by mechanisms independent of the enhanced stimulation of phospholipase C; amplification at the level of sequential protein kinase C and tyrosine kinase steps leads to the enhanced responsiveness of MAPK in the SHR derived cells.

Angiotensin II↗
PubMed1997

Activity and expression of Na(+)-H+ exchanger isoforms 1 and 3 in kidney proximal tubules of hypertensive rats.

Increased activity of the cellular Na(+)-H+ exchangers (NHEs), especially isoform 1 (NHE-1), is a recognized intermediate phenotype of hypertension. NHE activity has been demonstrated to be increased in proximal tubules of the spontaneously hypertensive rat (SHR). However, with the recent cloning of other members of this family of transporters, it is unclear which isoforms may contribute to this increased activity. We have used specific antibodies raised against glutathione-S-transferase fusion proteins of rat NHE-1 and NHE-3 to determine the relative contributions of these isoforms to the NHE activity in freshly isolated and cultured proximal tubule cells from SHR and Wistar-Kyoto (WKY) normotensive control rats. In freshly isolated proximal tubule cells, NHE activity was elevated almost 3-fold in SHR cells (P < .001), and in both rat strains, the contribution from NHE-1 and NHE-3 was approximately equal. Western blots of membranes from these cells showed equal amounts of NHE-1 protein in SHR and WKY cells. However, NHE-3 protein expression was increased 50% in SHR cells (P < .001), and this may account for the elevated activity of this isoform in SHR. The effect of culturing these cells in vitro was then examined. Although total NHE activity in both cell types was decreased during culture, this was mainly due to loss of expression of NHE-3 protein. NHE-1 activity was persistently elevated in the SHR cells in culture. These findings suggest that elevated NHE activity in SHR proximal tubules could be mediated by two mechanisms: (1) increased NHE-1 activity without any increased NHE-1 protein content that persists despite culture and may resemble those changes described for extrarenal tissues and (2) increased NHE-3 activity due to increased expression of NHE-3 protein. Disappearance of NHE-3 during culture implies that our culture conditions did not replicate the in vivo environment and may have removed the factors contributing to the increased NHE-3 expression in SHR cells.

Animals↗
PubMed1997

Disordered water homeostasis in Asian patients with schizophrenia.

OBJECTIVE: The aim of this study was to determine the prevalence of polydipsia-hyponatremia among patients with schizophrenia in an Asian mental hospital. METHOD: Seven hundred and twenty-eight inpatients with schizophrenia were assessed for polydipsia-hyponatremia using case notes reviews, specific gravity of urine, normalised diurnal weight gain, and serum sodium levels. RESULTS: One hundred and three (13.8%) patients had polydipsia, 30 (4.1%) had polydipsia-hyponatremia and 14 (1.9%) had a history of water intoxication. Eight of the 30 patients were receiving carbamazepine, three were on tricyclic antidepressants and two had diabetes mellitus and were on sulfonylureas. CONCLUSION: The prevalence of water intoxication among polydipsic patients was low compared to Western studies. This could be due to different methods of assessing polyuria, or ethnic differences and/or the prohibition of smoking in our patients. Certain medications might have also contributed to hyponatremia.

Adult↗
PubMed1996

Stimulated mitogen-activated protein kinase is necessary but not sufficient for the mitogenic response to angiotensin II. A role for phospholipase D.

Activation of the mitogen-activated protein kinase (MAPK) cascade has been widely associated with cell proliferation; previous studies have shown that angiotensin II (AII), acting on 7-transmembrane G protein-coupled receptors, stimulates the MAPK pathway. In this report we investigate whether the MAPK pathway is required for the mitogenic response to AII stimulation of vascular smooth muscle cells derived from the hypertensive rat (SHR-VSM). AII stimulates the phosphorylation of MAPK, as determined by Western blot specific for the tyrosine 204 phosphorylated form of the protein. This MAPK phosphorylation was inhibited by the presence of the inhibitor of MAPK kinase activation, PD 098059. Using a peptide kinase assay shown to measure the p42 and p44 isoforms of MAPK, the stimulated response to AII was inhibited by PD 098059 with an IC50 of 15.6 +/- 1.6 microM. The AII stimulation of [3H]thymidine incorporation was inhibited by PD 098059 with an IC50 of 17.8 +/- 3.1 microM. PD 098059 had no effect on AII-stimulated phospholipase C or phospholipase D (PLD) activity. When the SHR-VSM cells were stimulated with phorbol ester, there was an activation of MAPK similar in size and duration to the response to AII, but there was no significant enhancement of [3H]thymidine incorporation. There was also no activation of PLD by phorbol ester, while AII produced a robust PLD response. Diversion of the product of the PLD reaction by 1-butanol caused a partial loss of the [3H]thymidine response; this did not occur with tertiary butanol, which did not interfere with the PLD reaction. These results show that in these cells the MAPK cascade is required but not sufficient for the mitogenic response to AII, and suggest that the full mitogenic response requires both MAPK in conjunction with other signaling components, one of which is PLD.

Angiotensin II↗
PubMed1996

Glucose-induced changes in activity and phosphorylation of the Na+/H+ exchanger, NHE-1, in vascular myocytes from Wistar-Kyoto and spontaneously hypertensive rats.

Increased Na+/H+ exchanger (NHE) activity has been demonstrated in cells from patients with hypertension and diabetic nephropathy. Vascular myocytes from the spontaneously hypertensive rat (SHR) also exhibit increased NHE activity as compared with cells from the normotensive Wistar Kyoto rat (WKY). The interaction of increased glucose concentrations with NHE activity is unclear. The effect of glucose on NHE activity, NHE-1 (isoform 1) protein expression, and phosphorylation of cultured vascular myocytes from these rat strains was thus investigated. NHE activity was determined fluorometrically with 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF). A rabbit NHE-1-specific polyclonal antibody was used (1) to measure NHE-1 abundance in Western blots of cell extracts and (2) for immunoprecipitating 32P-labeled NHE-1. Cells from SHR exhibited increased NHE activity and NHE-1 phosphorylation as compared with cells from WKY, with similar NHE-1 protein content per cell. Incubation in 25 mmol.L-1 glucose for 24 hours led to increased NHE activity only in WKY cultures, with no change in NHE-1 protein but a concomitantly reduced NHE-1 phosphorylation. Changes in NHE activity in WKY cells were reversed by inhibition of protein kinase C. Incubation of SHR cells with 25 mmol.L-1 glucose did not enhance the increased NHE activity or NHE-1 phosphorylation present in these cells. Thus, high glucose levels have disparate effects on NHE activity and NHE-1 phosphorylation in cells from different rat strains. The glucose-induced increase in NHE-1 turnover number in WKY cells is not mediated by an increase in its direct phosphorylation, but is dependent on protein kinase C.

Animals↗
PubMed1996

Phorbol ester activation of the rat vascular myocyte Na(+)-H(+) exchanger isoform 1.

Vascular myocytes from the spontaneously hypertensive rat (SHR) demonstrate elevated Na(+)-H(+) exchanger activity associated with increased cell proliferation and hyperresponsiveness to agonists such as phorbol esters. Since the Na(+)-H(+) exchanger isoform 1 (NHE-1) is stimulated by protein kinase C, we have investigated the effects of phorbol esters on NHE-1 activity and its phosphorylation in vascular myocytes of these rats. SHR cells demonstrated a larger alkalinization response to 12-O-tetradecanoylphorbol 13-acetate than Wistar-Kyoto rat (WKY) cells. Kinetic analyses indicated that whereas 12-O-tetradecanoylphorbol 13-acetate increased the maximal transport capacity of NHE-1 in both cell types, affinity for H+ was increased in WKY cells and cooperativity for H+ at the internal modifier site was reduced in SHR cells. In neither cell type was the subcellular distribution of NHE-1 altered by phorbol ester stimulation. NHE-1 phosphorylation was markedly reduced in WKY cells stimulated by the phorbol ester, an effect abolished by inhibition of protein kinase C. In contrast, NHE-1 phosphorylation in quiescent SHR cells was approximately double that of WKY cells and was reduced after phorbol ester treatment. Inhibition of protein kinase C in SHR cells led to a marked elevation of NHE-1 phosphorylation that was not associated with a change in the exchanger activity, but WKY cells exhibited a small, insignificant rise in NHE-1 phosphorylation. Thus, the kinetic responses of NHE-1 to phorbol esters in vascular myocytes of these rat strains are different, the changes in exchanger kinetics of SHR resembling those described in human hypertension. NHE-1 phosphorylation has an inverse relationship with protein kinase C activity. However, modulation of NHE-1 phosphorylation may not be associated with concurrent alterations in activity, indicating a role for non-phosphorylation-dependent mechanisms.

Animals↗
PubMed1996

Angiotensin II-stimulated phospholipase C responses of two vascular smooth muscle-derived cell lines. Role of cyclic GMP.

Vascular smooth muscle cells of the spontaneously hypertensive rat (SHR) are known to show increased responsiveness to angiotensin II (Ang II) compared with cells of normotensive control Wistar-Kyoto rats (WKY). We investigated the hypothesis that differential levels of cGMP lead to the different responsiveness of the cells, using vascular smooth muscle cells in culture. cGMP levels in extracts of SHR-derived cells were lower than those of WKY-derived cells. This was true for both unstimulated cells and cells treated with equal concentrations of either sodium nitroprusside or S-nitroso-N-acetylpenicillamine. Stimulation of cells with Ang II did not affect levels of cGMP but increased levels of inositol 1, 4, 5-trisphosphate (IP3) and Ca2+, which were greater in SHR- than in WKY-derived cells. When SHR and WKY cells were preincubated with different concentrations of S-nitroso-N-acetylpenicillamine to generate similar cGMP levels in each cell type, the subsequent IP3 response to Ang II was the same in the two cell types. To reduce any influence of cGMP on responses, we permeabilized the cells with alpha-toxin. Stimulation of alpha-toxin-permeabilized the cells with high Ca2+ revealed an IP3 response in SHR- but not WKY-derived cells. Similarly, permeabilized SHR cells responded to Ang II but WKY cells did not. However, GTP and GTP gamma S elevated IP3 in both cell types. Taken together, these results indicate that the low response of WKY cells can be accounted for by the inhibitory influence of cGMP. However, when this inhibition is removed by permeabilization, further differences between the cells are revealed that will contribute to the elevated SHR response.

Angiotensin II↗
PubMed1995

Preparation and properties of nido-carborane-specific monoclonal antibodies for potential use in boron neutron capture therapy for cancer.

As the first step of a research program aimed at developing a bispecific monoclonal antibody system for the delivery of boron-rich molecules to tumor cells for boron neutron capture therapy, monoclonal antibodies (mAbs) were produced against an anionic nido-carborane derivative, 4-[7,8-dicarbadodecahydroundecaborat(-1)-7-yl]butanoic acid. Two IgG subclass mAbs, designated HAW101 and HAW102, were identified that specifically bound the anionic nido-carborane hapten, as well as a variety of other anionic nido-carborane cage derivatives. By using surface plasmon resonance technology, the affinity constants of HAW101 and HAW102 were determined to be 1.9 x 10(9) and 6.8 x 10(8) M-1, respectively. A diverse array of 7-substituted and 7,8-disubstituted anionic nido-carborane derivatives reacted with the mAb HAW101 in competition ELISA, whereas anionic closo-polyhedral boranes showed negligible binding, suggesting a role for the open nido-carborane cage structure. These results suggest that mAbs such as HAW101, which bind anionic nido-carboranes, are useful in the development of bispecific mAbs for specific targeting and enhanced boron delivery to tumor sites.

Antibodies, Monoclonal↗
PubMed1995

Culture density and the activity, abundance and phosphorylation of the Na+/H+ exchanger isoform 1 in human fibroblasts.

The Na+/H+ exchanger isoform 1 (NHE-1) is a ubiquitous membrane glycoprotein present on most eukaryotic cells. Its activity, abundance and phosphorylation are regulated by a variety of growth factors and agonists. Although cell contact and inhibition of proliferation may reduce its activity, little is known of the influence of cell culture density on these measurements. The effect of culture density on the intracellular pH (pHi) and activity of NHE-1 of human MRC5 fibroblasts was thus investigated using fluorometry with BCECF, NHE-1 abundance with western blotting and NHE-1 phosphorylation using specific polyclonal antibodies. Proliferating cells in low density cultures had lower pHi and NHE-1 activity (per litre of cell water) than contact inhibited confluent cultures. Such cells in low density cultures were larger than those in very confluent cultures. NHE-1 activity per cell and NHE-1 protein abundance also showed an increasing trend with culture density. However, the turnover number of NHE-1 remained at around 3000 s-1 in low density and sub-confluent cultures, only decreasing in very confluent cultures. Moreover, NHE-1 phosphorylation declined with increased culture density. Cell culture density has profound effects on NHE-1 activity, abundance and turnover number, with associated changes in NHE-1 phosphorylation.

Cell Line↗
PubMed1995

Na+/H+ exchanger isoform-1 abundance in skin fibroblasts of type I diabetic patients with nephropathy.

In diabetic nephropathy and essential hypertension, the cellular Na+/H+ exchanger (NHE) exhibits increased activity. Whether this reflects increased numbers of NHE isoform-1 (NHE-1) transporters or increased turnover per molecule has not been established. We have used a specific polyclonal antibody directed toward the C-terminal of NHE-1 to measure NHE-1 content in cultured skin fibroblasts from diabetic patients with (DN) and without (DCON) nephropathy and normal controls (CON). NHE-1 content in fibroblasts from DN subjects was significantly less than that in the other two groups. This suggests that increased NHE activity in diabetic nephropathy is attributed to increased NHE-1 turnover per site rather than increased NHE-1 expression.

Adult↗
PubMed1995

Effect of extracellular potassium concentration on the sodium-potassium pump rate in human lymphocytes.

1. The purpose of this study was to determine whether physiological changes in extracellular free [K+] cause significant changes in the Na(+)-K+ pump rate and intracellular free [Na+]. 2. The Na(+)-K+ pump rate was measured in human lymphocytes by determining ouabain-sensitive 86Rb+ influx at several concentrations of K+. The Na(+)-K+ pump rate increased within the physiological range of extracellular free [K+] (K1/2 = 1.5 mmol/l). 3. To test the hypothesis that elevation of extracellular free [K+] reduces intracellular free [Na+] rapidly, which in turn then slows the pump rate during experimental incubations, lymphocyte intracellular free [Na+] was measured using the fluorochrome sodium-binding benzofuran isophthalate. With larger elevations of extracellular free [K+], intracellular free [Na+] dropped more rapidly. Thus previous discrepancies among determinations of K1/2 may be the result of variations in incubation times, which can skew the pump rates measured during incubations in various extracellular free [K+] values. Steady-state intracellular free [Na+] varied inversely with extracellular free [K+].

Adult↗
PubMed1995

Na(+)-H+ antiporter phenotype, abundance, and phosphorylation of immortalized lymphoblasts from humans with hypertension.

Previous studies have demonstrated an elevated Na(+)-H+ exchanger activity in various cell types from patients with essential hypertension. The phenotype of an increased maximal transport capacity is preserved in Epstein-Barr virus immortalized lymphoblasts from hypertensive patients. The mechanisms underlying this abnormality are unclear. In this study, we used lymphoblasts from hypertensive patients and normotensive control subjects with and without a family history of hypertension to determine (1) Na(+)-H+ exchanger activity using fluorometry with the pH indicator 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein, (2) Na(+)-H+ exchanger isoform 1 abundance with specific polyclonal antibodies, and (3) Na(+)-H+ exchanger phosphorylation by immunoprecipitation of the 32P-labeled transporter. Na(+)-H+ exchanger activity (in millimoles per liter per minute) measured when pHi was clamped at 6.0 was significantly higher in cells from hypertensive patients (18.8 +/- 0.6, P < .001) and those subjects with a family history of hypertension (16.4 +/- 0.6, P < .001) compared with normotensive control subjects (12.9 +/- 0.6). Exchanger abundance was identical in all three groups of subjects, indicating that increased activity in the hypertensive group was due to an elevated turnover number of the exchanger. Na(+)-H+ exchanger phosphorylation in quiescent cells was significantly elevated in cells from hypertensive patients (1.58 +/- 0.16, P < .001) compared with control subjects (1.00 +/- 0.07), and cells from normotensive subjects with a hypertensive family history showed intermediate values (1.23 +/- 0.14). Identical changes in Na(+)-H+ exchanger function and phosphorylation have been demonstrated in vascular smooth muscle cells from spontaneously hypertensive rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1995

Na(+)-H+ exchanger isoform 1 phosphorylation in normal Wistar-Kyoto and spontaneously hypertensive rats.

Increased activity of the cellular Na(+)-H+ exchanger (NHE) has been documented in various cell types in essential hypertension and in vascular myocytes of the spontaneously hypertensive rat (SHR). The mechanism underlying this abnormality is unclear. Because the NHE can be activated by phosphorylation, we examined phosphorylation of the Na(+)-H+ exchanger isoform 1 (NHE-1) as one possible mechanism for its increased turnover number in cultured vascular myocytes of the SHR. A polyclonal rabbit antibody against a fusion protein consisting of beta-galactosidase and the C-terminus of NHE-1 was used to immunoprecipitate 32P-labeled NHE-1 from cell extracts of SHR and Wistar-Kyoto (WKY) rat vascular myocytes in the absence and presence of 10% fetal calf serum. Immunoprecipitates were separated by SDS-PAGE, and 32P-labeled NHE-1 was quantified from autoradiographs. Similar amounts of NHE-1 protein were detected on Western blots of the cultured vascular myocytes from SHR and WKY rats. In quiescent cells, NHE-1 was significantly more phosphorylated in SHR myocytes than in WKY myocytes (2.17 +/- 0.06-fold enhancement [mean +/- SEM]; P < .001, n = 8). The addition of fetal calf serum to quiescent cells had no significant effect on the phosphorylation of NHE-1 in SHR myocytes. However, NHE-1 phosphorylation fell transiently in serum-treated WKY myocytes, with recovery to control levels after 20 minutes. Measurement of NHE activity using fluorometry confirmed elevated activity in the quiescent SHR myocytes compared with WKY myocytes. Fetal calf serum led to further enhancement of NHE activity in both cell types. These findings suggest that the increased NHE activity in quiescent SHR myocytes may be correlated with enhanced NHE-1 phosphorylation and that serum stimulates NHE activity in both cell types without a further increase in total NHE-1 phosphorylation, indicating a role for non-phosphorylation-dependent regulatory mechanisms.

Animals↗
PubMed1995

Phosphorylation and activity of Na+/H+ exchanger isoform 1 of immortalized lymphoblasts in diabetic nephropathy.

In both essential hypertension and diabetic nephropathy (DN), the ubiquitous cellular Na+/H+ exchanger (NHE) exhibits altered kinetics with increased transport activity. The mechanism for this phenotype and its dependence on the presence of serum are unknown, but increased lymphoblast NHE activity in DN has been attributed to a defect in post-translational processing of NHE-1 rather than an increased cellular exchanger number. Phosphorylation of NHE-1 has been proposed to play a role in its activation in a variety of cell models. We have examined, therefore, the role of NHE-1 phosphorylation and the effect of serum in determining the increased NHE-1 activity in lymphoblasts from patients with DN. Cells from these patients exhibited increased NHE activity in the presence and absence of fetal calf serum (range 42-59%, P < 0.005, analysis of variance) and an increased proliferation rate (P < 0.01) when compared with cells from both normoalbuminuric diabetic patients and non-diabetic control subjects. However, NHE-1 abundance was very similar among all groups in the presence and absence of serum, indicating that increased NHE activity in cells of nephropathy patients was due to an increased turnover number. This nephropathy phenotype was not accompanied by an increased net phosphorylation of NHE-1 in the presence or absence of serum. Our findings suggest that increased NHE-1 activity in cells of DN patients is independent of the presence of serum and is not attributable to altered NHE-1 phosphorylation. Additional post-translational mechanisms for activation of NHE-1, therefore, may be involved.

Adult↗