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N Shukla

Publications and source records attributed to N Shukla.

At least 19 recordsLinked to original sources
PubMed2006

Reactive oxygen species and erectile dysfunction: possible role of NADPH oxidase.

Erectile dysfunction (ED) is a widespread condition, the incidence of which is increasing globally. ED is also indicative of underlying vasculopathy and represents a predictor of more serious cardiovascular disorders. Understanding the aetiology of ED may therefore provide invaluable pointers to the pathobiology of other cardiovascular diseases (CVDs) and syndromes. It follows, too, that therapeutic interventions that are successful in treating ED may, ipso facto, be effective in treating the early stages of conditions that include atherosclerosis, angina, plaque rupture and diabetic angiopathy. One common pathological denominator in both CVD and ED is oxidative stress, that is, the overproduction of reactive oxygen species (ROS), in particular, superoxide (O(2)(*-)) and hydrogen peroxide (H(2)O(2)). In this review, therefore, we consider the aetiology and pathobiology of O(2)(*-) in promoting ED and focus on NADPH oxidase as an inducible source of O(2)(*-) and H(2)O(2). Therapeutic strategies aimed at reducing oxidative stress to improve erectile function are also discussed.

Animals↗
PubMed2005

Acute hypoxia simultaneously induces the expression of gp91phox and endothelial nitric oxide synthase in the porcine pulmonary artery.

BACKGROUND: The effect of hypoxia on the formation of superoxide (O2-), the expression of gp91phox and endothelial NO synthase (eNOS) were studied in pig intact pulmonary artery (PA) segments and PA vascular smooth muscle cells (PAVSMCs) and PA endothelial cells (PAECs). METHODS: Segments and cells were incubated under hypoxic conditions for 2 hours (with or without enzyme inhibitors) and the formation of O2- measured spectrophotometrically. Protein expression was assessed using Western blotting and immunocytochemistry. RESULTS: Hypoxia promoted the formation of O2- in PA segments, PAVSMCs and PAECs, an effect inhibited by diphenylene iodonium and apocynin (NAD[P]H oxidase inhibitors). Hypoxia induced O2- formation was enhanced by inhibition of eNOS and augmented by endotoxin and cytokines and re-oxygenation. Hypoxia also promoted the expression of gp91phox and eNOS. In intact PA segments hypoxia induced the expression of nitrotyrosine and eNOS in the endothelium. CONCLUSIONS: The simultaneous upregulation of NAD[P]H oxidase and eNOS in response to hypoxia in the PA results in the simultaneous formation of O2-, NO, and ONOO-. This may represent either a protective mechanism designed to counter the pro-oxidant effect of hypoxia or a novel pathological mechanism underlying the progression of acute respiratory distress syndrome (ARDS).

Animals↗
PubMed2004

Low micromolar concentrations of copper augment the impairment of endothelium-dependent relaxation of aortae from diabetic rabbits.

Both diabetes mellitus (DM) and elevated plasma copper concentrations are risk factors for cardiovascular disease (CVD). DM is associated with impaired endothelial nitric oxide (NO) and with excess superoxide (O2*-) formation. Copper is also elevated in DM and is also associated with the generation of O2*-. To explore possible interactions between DM and copper, the effect of exogenous copper (CuCl2) on endothelium-dependent relaxation and cyclic guanosine monophosphate (GMP) formation was investigated in aortae from diabetic rabbits. Rabbits were rendered diabetic by intravenous injection of alloxan. Six months after induction of DM, the aortae were excised, cut into rings, and mounted in an organ bath for isometric measurement of acetylcholine (Ach)-evoked relaxation in rings precontracted with phenylephrine (PE). In parallel studies, cyclic (c)GMP formation by aortic rings following stimulation with Ach, calcium ionophore A23187 (A23187) and sodium nitroprusside (SNP) was assessed using radioimmunoassay. The effect of copper on these parameters was then studied using the same methods. Ach-evoked relaxation and Ach- and A23187-evoked cGMP formation were significantly impaired in aortae from diabetic rabbits compared to controls, effects that were reversed with superoxide dismutase (SOD) and catalase (CAT). In contrast, there were no significant differences in SNP-stimulated relaxation or cGMP formation in aortae from diabetic rabbits compared to controls. Copper (1 to 10 micromol/L) promoted a further significant inhibition of Ach-stimulated relaxation in aortae from diabetic but not control rabbits. This reduction by copper was again reversed by SOD and CAT. We conclude that copper augments the reduction of NO bioavailability, which is already impaired in aortae from diabetic rabbits due to excess production of O2*- and H2O2. These results indicate that patients with DM may be susceptible to copper-mediated vasculopathy at much lower concentrations than those that promote vasculopathy in nondiabetic patients.

Acetylcholine↗
PubMed2003

Role of the endothelium and nitric oxide synthases in modulating superoxide formation induced by endotoxin and cytokines in porcine pulmonary arteries.

BACKGROUND: The interactive roles of cytokines, endotoxins, superoxide (O(2)(*-) ) and nitric oxide (NO) in the pathogenesis of adult respiratory distress syndrome (ARDS) have not been fully elucidated. The effects of tumour necrosis factor-alpha (TNF-alpha), interleukin 1alpha (IL-1alpha), and lipopolysaccharide (LPS) and the role of NO and the endothelium in mediating O(2)(*-) formation were therefore investigated in intact porcine pulmonary arteries in vitro. METHODS: Intrapulmonary artery (PA) segments were obtained from White Landrace pigs (25-35 kg) and incubated with LPS, IL-1alpha, and TNF-alpha and O(2)(*-) release was measured by the superoxide dismutase (SOD) inhibitable reduction of ferricytochrome c. The source of O(2)(*-) formation was determined using a number of enzyme inhibitors. The role of NO was explored using NO synthase (NOS) inhibitors and the distribution of NOS isoforms and peroxynitrite (ONOO(-), an index of NO-O(2)(*-) interactions) assessed by immunocytochemistry. RESULTS: LPS, IL-1alpha, and TNF-alpha promoted the formation of O(2)(*-) from PA compared with untreated controls in a time and dose dependent manner, an effect markedly enhanced by removal of the endothelium but completely inhibited by the NADPH oxidase inhibitor diphenylene iodonium chloride (DPI). L-NAME and the eNOS inhibitor N(5)-(1-iminoethyl)-ornithine (L-NIO) enhanced O(2)(*-) formation from PA (with endothelium) in response to IL-1alpha and TNF-alpha but had no effect on LPS mediated O(2)(*-) formation, whereas L-NAME and the iNOS inhibitor L-N(6)-(1-iminoethyl)-lysine-HCl (L-NIL) enhanced O(2)(*-) formation only in response to LPS. CONCLUSIONS: LPS, IL-1alpha, and TNF-alpha promote O(2)(*-) formation through an upregulation of NADPH oxidase activity which is augmented by removal of the endothelium, as well as the inhibition of eNOS (in the case of cytokines) and iNOS (in the case of LPS). The concomitant expression of NOS isoforms (and NO formation) with that of NADPH oxidase may therefore constitute a protective system designed to remove O(2)(*-) through the formation of ONOO(-). If this is so, the integrity of the endothelium may be axiomatic in the progression and severity of ARDS.

Animals↗
PubMed2002

Homocysteine enhances impairment of endothelium-dependent relaxation and guanosine cyclic monophosphate formation in aortae from diabetic rabbits.

AIMS/HYPOTHESIS: Both diabetes mellitus and hyperhomocysteinaemia are risk factors for cardiovascular disease and are associated with impaired endothelial nitric oxide and with excess superoxide formation. To explore potential vasculopathic interactions between these risk factors, the effect of homocysteine on endothelium-dependent relaxation and cyclic GMP formation was investigated in aortae from diabetic rabbits. METHODS: Rabbits were rendered diabetic by intravenous injection of alloxan. Six months later, the aortae were excised, cut into rings and mounted in an organ bath for isometric measurement of acetylcholine-evoked relaxation in rings pre-contracted with phenylephrine. Cyclic GMP formation by aortic rings after stimulation with acetylcholine, calcium ionophore A23187 and sodium nitroprusside was assessed using radioimmunoassay. The effect of homocysteine on these parameters was then studied. RESULTS: Ach-evoked relaxation and cyclic GMP formation induced with acetylcholine and calcium ionophore A23187 were impaired in aortae from diabetic rabbits compared with the control rabbits, effects that were reversed with superoxide dimutase (SOD) and augmented by 10-100 micro mol/l homocysteine, an effect again reversed by SOD. CONCLUSION/INTERPRETATION: These data show that the bioavailability of nitric oxide is reduced in aortae from diabetic rabbits due to excess production of superoxide, an effect augmented by homocysteine. These results indicate that patients with diabetes mellitus could be susceptible to homocysteine-mediated angiopathy at lower concentrations than those that promote vasculopathy in non-diabetic patients.

Acetylcholine↗
PubMed2001

Thapsigargin inhibits angiogenesis in the rat isolated aorta: studies on the role of intracellular calcium pools.

OBJECTIVE: Since the role of Ca2+ in angiogenesis is not fully understood, we investigated the effect of thapsigargin (TG: depletes intracellular Ca2+ pools) and other Ca2+ modulators [ionomycin, calcium ionophore A23187 and dibutyrylhydroquinone (DBHQ)] on in vitro angiogenesis by rat aortic rings. METHODS: Aortae from Sprague-Dawley rats were cut into 2-mm rings, embedded in a fibrin clot and cultured for 15 days in serum-free medium containing drugs and the microvessels counted. Rings were also pre-treated with TG and Ca2+ modulators for 1 h prior to embedding and culture. Viability was examined by the measurement of lactic acid dehydrogenase release. Rings were also treated with hydrocortisone and lavendustin A (a tyrosine kinase inhibitor), as positive controls. The effect of TG on the proliferation and migration of human umbilical artery endothelial cells (HUVECs) was studied in parallel. RESULTS: TG significantly inhibited microvessel formation and HUVEC proliferation and migration in a dose-dependent manner, all at <10 nmol/l, without affecting viability. In contrast, ionomycin, A23187 and DBHQ were cytotoxic at inhibitory concentrations. Continual exposure to hydrocortisone and lavendusin A also inhibited angiogenesis without affecting viability. CONCLUSION: Since low concentrations of TG deplete intracellular Ca2+ stores, it is concluded that these pools play a central role in mediating angiogenesis.

Analysis of Variance↗
PubMed2000

Inhibition of diabetic bladder smooth muscle cell proliferation by endothelin receptor antagonists.

Urinary bladder hypertrophy and hyperplasia are well recognised in diabetic cystopathy. The urinary bladder is known to synthesise endothelin-1 (ET-1), a potent vasoconstrictor peptide with mitogenic properties. Using diabetic New Zealand White (NZW) rabbits, we investigated the potential role of ET receptor subtypes (ETA and ET(B)) on the proliferation of bladder smooth muscle cells (SMC). Diabetes mellitus was induced in adult male NZW rabbits. After 6 months, control (n = 6) and diabetic (n = 6) bladders were removed and SMC from the dome and bladder neck were grown using standard explant methodology. At passage two, the cells were made quiescent and then further incubated in foetal calf serum (FCS), control age-matched rabbit serum (CRS) or diabetic rabbit serum (DRS) in the presence or absence of ET(A)-antagonist (BQ123) or ET(B)-antagonist (BQ788). SMC proliferation was then measured with 5-bromo-2'deoxy-uracil 24 h later and by cell counting (using a haemocytometer) at 48 h. Neither BQ123 nor BQ788 influenced detrusor or bladder neck SMC proliferation in FCS or CRS. However, in the presence of DRS, BQ123 and BQ788 significantly inhibited diabetic detrusor and bladder neck SMC proliferation at 30 and 100 nmol/l (P < 0.03 and P < 0.01, respectively). Cell counts were also significantly reduced from the diabetic detrusor and bladder neck (P < 0.01 and P < 0.03 with BQ123 and BQ788, respectively). These results suggest that ET may play a pathophysiological role in the bladder SMC hyperplasia associated with diabetes mellitus.

Animals↗
PubMed2000

Possible role of endothelin-1 in the rabbit urinary bladder hyperplasia secondary to partial bladder outlet obstruction.

OBJECTIVES: Urinary bladder hypertrophy and hyperplasia are common features of bladder outlet obstruction (BOO). The urinary bladder is known to synthesize endothelin-1 (ET-1), which is a potent vasoconstrictor peptide with mitogenic properties. Using an animal model of partial BOO, we investigated the potential role of ET-1 and its receptor subtypes (ET(A) and ET(B)) in bladder smooth muscle cell (SMC) proliferation. MATERIALS AND METHODS: Partial BOO was produced in adult male New Zealand White rabbits. After 3 weeks, the bladder was removed and SMCs from the dome and bladder neck were grown using standard explant methodology. At passage 2, the cells were made quiescent and then further incubated in foetal calf serum (FCS), control age-matched rabbit serum (CRS) or partial BOO serum (BRS) in the presence or absence of ET(A)-antagonist (BQ123) or ET(B)-antagonist (BQ788). SMC proliferation was then measured 24 h later with 5-bromo-2'deoxy-uracil and by cell counting using a haemocytometer at 48 h. Immunostaining for alpha-actin was performed on detrusor and bladder neck cells to confirm the presence of smooth muscle cells. RESULTS: BQ123 and BQ788 did not influence detrusor or bladder neck SMC proliferation in FCS or CRS. However, in the presence of BRS, BQ123 and BQ788 (100 nmol/L) significantly (p = 0.008) inhibited detrusor and bladder neck SMC proliferation. Cell counts were significantly reduced from the detrusor (p = 0.03, p = 0.01 with BQ123 and BQ788, respectively) and bladder neck (p = 0.01 for both BQ123 and BQ78). CONCLUSIONS: These results suggest that ET antagonists may have a role in preventing SMC hyperplasia associated with partial BOO.

Animals↗
PubMed2000

Endothelin-1 and urinary bladder hyperplasia following partial bladder outlet obstruction.

Urinary bladder hypertrophy and hyperplasia is a common feature of bladder outlet obstruction (BOO). The urinary bladder is known to synthesize endothelin-1 (ET-1). ET-1 is a potent vasoconstrictor peptide with mitogenic properties. Using an animal model of partial BOO we investigated the potential role of ET-1 and its receptor subtypes [endothelin-A and -B (ET(A) and ET(B))] in bladder vascular smooth muscle cells (SMC) proliferation. In the presence of 3-week-old BOO serum, ET(A) and ET(B) antagonists significantly (p = 0.008) inhibited detrusor and bladder neck SMC proliferation. Cell counts were significantly reduced from the detrusor (p = 0.03, p = 0.01 with ET(A) and ET(B) antagonists, respectively) and bladder neck (p = 0.01 for both ET(A) and ET(B) antagonists). These results suggest that ET-1 antagonists may prevent SMC hyperplasia associated with partial BOO.

Animals↗
PubMed1999

Anti-heat-shock protein 70 kDa antibodies in vascular patients.

INTRODUCTION AND AIM OF STUDY: there is recent evidence that the immune system plays an essential role in the pathogenesis of atherosclerosis, with both cellular and humoral mechanisms being involved. Heat-shock proteins (HSPs) have been detected in atherosclerotic lesions, and antibodies to HSPs have also been found to be raised in patients with carotid stenoses. The aim of our study was to examine the level of anti-HSP70 antibodies in patients with other vascular diseases. MATERIALS AND METHODS: a questionnaire was designed for the subjects in the study, with documentation of clinical details and ankle-brachial pressure index. Patients with concomitant infection, malignancy, hepatorenal failure, or recent surgery were excluded. Enzyme-linked immunosorbent assay (ELISA) was used to identify anti-HSP70 antibodies in the sera in different dilutions. Graphs of optical density (OD) vs. negative log dilution were plotted, the gradient of which was taken to be the estimated optical density for each subject (proportional to antibody level). Our groups consisted of controls (n =21, mean age 59.0+/-19.2), lower limb claudicants ( n =19, mean age 60.0+/-12.6), patients with lower-limb critical ischaemia ( n =22, mean age 68.5+/-10.07), and patients with abdominal aortic aneurysms ( n =20, mean age 69.9+/-6.2). RESULTS: we found no correlation between age and the estimated OD in our subjects (Spearman's correlation coefficient ( r )=0.123, one-tailed p value was 0.135). Patients with intermittent claudication, critical lower limb ischaemia, and aneurysms had higher estimated OD, and therefore higher anti-HSP70 antibody levels, than controls (Mann-Whitney test p =0.0127, 0.0037, 0.0008, respectively). CONCLUSIONS: our data provide the first evidence of a correlation between anti-HSP70 antibodies and different types of vascular diseases, suggesting that HSP70 might be involved in the pathogenesis and propagation of atherosclerosis. Since the immune response to HSPs can be modulated, this opens up the possibility of new therapeutic approaches.

Adult↗
PubMed1999

Inhibition of intracellular Ca2+ mobilisation by low antiproliferative concentrations of thapsigargin in human vascular smooth-muscle cells.

Low nanomolar concentrations of thapsigargin, a modulator of intracellular Ca2+ ([Ca2+]i) pools, inhibit vascular smooth-muscle cell (VSMC) proliferation. Because the mechanisms underlying this effect have not been defined, the effect of antiproliferative concentrations of thapsigargin on [Ca2+]i in fura-2-loaded VSMCs was studied by using dynamic video imaging of [Ca2+]i. After seeding on coverslips, human VSMCs were incubated for 1-48 h with thapsigargin before loading with fura-2 or during imaging. Mobilisation of [Ca2+]i was stimulated with 1 microM ionomycin in Ca2+-free medium and the increase in [Ca2+]i detected by using Ca2+ imaging techniques. Continuous exposure of cells to low concentrations of thapsigargin (which failed measurably to increase in [Ca2+]i) reduced the ionomycin response in a time- and dose-dependent manner (100% inhibition at 10 nM thapsigargin after 1 h exposure). After exposure of cells to 10 nM thapsigargin for 1 h followed by washing and further incubation for < or = 72 h, there was a time-dependent recovery of the ionomycin response. Because the concentrations of thapsigargin and exposure times are identical to those that inhibit replication in VSMCs, it is proposed that depletion of [Ca2+]i pools mediates the inhibitory effect of thapsigargin on VSMC proliferation.

Calcium↗
PubMed1998

Nitric oxide, prostacyclin and cyclic nucleotide formation in externally stented porcine vein grafts.

Non-restrictive, porous, external stents inhibit neointima formation in porcine vein grafts. Since the mechanisms underlying these effects are unknown we investigated the impact of this external stent on factors known to inhibit vascular smooth muscle cell proliferation: prostacyclin (PGI2), nitric oxide (NO), cAMP and cGMP formation in different regions of stented and unstented porcine vein grafts. Paired stented and unstented saphenous vein-carotid artery interposition grafting was carried out in Landrace pigs. One month after surgery, the vessels were excised and the formation of PGI2, cAMP and cGMP determined using radioimmunoassay and nitric oxide synthase (NOS) distribution studied using autoradiography and histochemistry. There were no significant differences between PGI2, cAMP and cGMP (nitroprusside-stimulated) formation in the medial/intimal regions of grafts of stented vein graft and ungrafted saphenous vein whereas all were significantly reduced in unstented vein graft. A23187-stimulated cGMP formation (mediated by NO release) and NOS content was significantly greater in the medial/intimal region of stented and unstented vein graft compared to ungrafted saphenous vein, indicating induction of endothelial NOS (eNOS) in both types of graft. This normalisation of the PGI2-cAMP axis and guanylyl cyclase activity in the medial/intimal region may contribute to the beneficial impact of the external stent on vein graft thickening. The increase in eNOS in both stented and unstented vein grafts mitigates against this isoform as playing a role in mediating the inhibitory effect of the stent on neointima formation. In the adventitia of both stented and unstented grafts there was an increase in PGI2, cAMP and cGMP formation compared to ungrafted saphenous vein, the production being greater in the stented compared to the unstented graft. In the adventitia of stented veini grafts, NOS, detected with NAPDH diaphorase staining, was associated with microvessels as well as with inflammatory cells. Taken together, these data are suggestive of a role for PGI2 and NO in promoting microangiogenesis in the adventitia of stented vein grafts which may in turn minimize graft hypoxia, an established contributory factor to neointima formation.

Animals↗
PubMed1998

Role of female sex steroids in regulating cholesteryl ester transfer protein in transgenic mice.

The role of sex steroids in the regulation of cholesteryl ester transfer protein (CETP) was examined in the following groups of female transgenic mice carrying the human CETP gene: (1) normal, (2) ovariectomized, (3) ovariectomized and treated with estrogen; (4) ovariectomized and treated with progesterone; (5) ovariectomized and treated with both hormones, and (6) ovariectomized and treated with tamoxifen. CETP activity was measured in the plasma, and in the particulate and the soluble fractions of liver, muscle, and adipose tissue. Human CETP specific activity was determined by taking the difference of cholesterol ester transfer in the presence and absence of an antibody (TP2) against human CETP Ovariectomy reduced hormone levels, but did not completely abolish them from the circulation. Plasma CETP activity was significantly reduced in the tamoxifen group. There were significant reductions in CETP in liver homogenate and the soluble fraction, as well as in the particulate fraction of adipose with ovariectomy. Hormone replacement did not restore CETP activity in either the plasma or the tissues. Tamoxifin treatment resulted in a decrease in CETP activity in both fractions of liver, but had no effect on adipose. In the soluble fraction of adipose tissue and both fractions of muscle, only trace CETP activity was detected. We conclude that (1) minimal amounts of sex steroid hormones may be sufficient to affect CETP expression; (2) the effects of sex steroid hormones vary among tissues; and (3) in addition to the sex steroids, factor(s) from the ovary are needed for the full expression of CETP in this animal model.

Animals↗
PubMed1997

Study of pulmonary functions in swimmers of Lucknow city.

In this study, the pulmonary functions of young swimmers of K.D. Singh Babu Stadium, Lucknow were studied, 20 swimmers of the age group of 15-20 years were chosen along with 15 students as controls. The parameters taken into account were Forced Vital Capacity (FVC), Forced Expiratory Volume (FEV1) and Peak Expiratory Flow Rate (PEFR). Two sets of readings were recorded. One before the start and the other at the end of the practice season. These readings were compared to each other and also with the values of the controls. The results indicate that swimming has considerable effect on enhancing lung functions of an individual as FVC, FEV1, PEFR were significantly raised in swimmers after swimming session. Such results suggest that swimming may be induced in the exercise programme for rehabilitation of respiratory patients who have compromised lung functions.

Adolescent↗
PubMed1997

Short-term exposure to low concentrations of thapsigargin inhibits replication of cultured human vascular smooth muscle cells.

BACKGROUND: Central to the pathophysiology of stenosis following balloon angioplasty and arterial bypass surgery is proliferation of vascular smooth muscle cells (VSMCs). To investigate the role of calcium (Ca2+) in VSMC proliferation, the effect of thapsigargin, Ca2+ ionophore A23187, ionomycin, cyclopiazonic acid and di-tert-butylhydroquinone (all of which raise intracellular Ca2+ levels) on the proliferation of cultured human VSMCs was observed. METHODS: Cultured VSMCs from human saphenous vein were treated with calcium-modulating drugs and proliferation was assessed by determining [3H]thymidine and 5-bromo-2'-deoxyuridine incorporation and cell number. RESULTS: Over a 48-h exposure, thapsigargin inhibited VSMC replication (median 50 per cent maximal inhibitory concentration 2 nmol/l) whereas the other drugs were much less effective. Short-term exposure (5, 10, 30 and 60 min) to thapsigargin elicited a significant dose-dependent inhibition of VSMC replication whereas, again, the other drugs were without significant effect. CONCLUSION: Thapsigargin-sensitive intracellular Ca2+ pools play a key role in controlling VSMC proliferation and specialized means of administering thapsigargin may constitute a possible approach to preventing stenosis.

Adult↗