Biomedical subjects
Publications and source records attributed to Dileep Kumar Rohra.
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BACKGROUND: In Pakistan, there is no mechanism to monitor the drug promotional campaign by pharmaceutical industry despite the fact that there is enough evidence that irrational pharmacotherapy is increasingly encountered even in the developed countries due to unethical practices of pharmaceutical promotion. Objectives. To audit the drug promotional claims made by the pharmaceutical companies in Pakistan. METHODS: Drug promotional pamphlets and brochures containing claims for the drugs, which were circulated by the pharmaceutical representatives were collected from 122 general practitioners (GPs) from Karachi and Larkana cities of the Sindh Province. The claims were critically analyzed and audited with the help of currently available evidence in the medical literature. RESULTS: 345 distinct advertisements covering 182 drugs from different manufacturers were critically analyzed for information content. Sixty two out of 345 (18%) of the reviewed advertisements were adjudged to be misleading / unjustifiable, which were again classified as, exaggerated (32%), ambiguous (21%), false (26%), and controversial (21%). The primary source of information (approximately 78%) about the newly launched drugs for the GPs was found to be the pharmaceutical representatives followed by hospital doctors (5%) and colleagues (5%). Furthermore, 110 (90%) GPs were of the view that the drug promotion has definitely an influence on their prescribing pattern. CONCLUSIONS: Since GPs in Pakistan rate pharmaceutical companies as their primary source of information regarding drugs, it can be anticipated that inappropriate advertisement claims would lead to irrational prescribing if physicians had no any other information to follow.
Metabolic acidosis is associated with various clinical situations including diabetes mellitus and renal diseases. The aim of this study was to investigate the effects of acidosis on the resting as well as precontracted human left internal mammary artery. The vessels were obtained from the patients undergoing coronary artery bypass grafting surgery at The Aga Khan University Hospital, Karachi. Left internal mammary artery was cut into rings and isometric tension recording experiments were performed. Decrease in pH of the bathing solution from 7.4 to 6.8 had no effect on the resting tension of left internal mammary artery, whereas, acidic pH markedly relaxed the contractions to 24.8 mM KCl and 300 nM phenylephrine. Interestingly, when the KCl- or phenylephrine-contracted rings were treated with 3 microM glibenclamide; an inhibitor of ATP-sensitive potassium (K(ATP)) channels, the relaxant effect of acidosis was abolished. Similarly, acidosis failed to cause relaxation of 100 nM endothelin-1-induced contraction in Ca2+-free bathing solution or in the presence of a voltage-dependent Ca2+ channel inhibitor, verapamil (10 microM), whereas, endothelin-1-induced contraction was attenuated by acidosis in Ca2+-containing normal solution. From all these data, it is concluded that under the acidic pH conditions, opening of K(ATP) channels occurs; resulting in the hyperpolarization, decrease in Ca2+ influx via voltage-dependent Ca2+ channels and subsequent relaxation of human left internal mammary artery.
This study was conducted to investigate the influence of extracellular Cl- ([Cl-]o) on the intracellular pH (pHi) regulation and the contractile state of the isolated aorta from Wistar Kyoto (WKY) rats. Isometric tension recording and fluorometry techniques were utilized to measure contractile response and pHi in isolated aortic strips. Decreasing extracellular pH (pHo) from 7.4 to 6.5 produced a marked contraction, which was 75.8 +/- 5.6% of the 64.8 mmol/l KCl-induced contraction. The acidosis-induced contraction was significantly attenuated in low [Cl-]o solution, the magnitude of which was 56.0 +/- 3.0% of the 64.8 mmol/l KCl-induced contraction. Decreasing pHo of the normal solution to 6.5 rapidly decreased pHi in aortic smooth muscle cells and produced a corresponding contraction. When the pHo was decreased in low [Cl-]o solution, a rapid fall in pHi followed by reversal of pHi changes, in a time-dependent manner was observed, despite low pHo. Omission of HCO3- from the low [Cl-]o solution restored the contractile response to acidosis, which was comparable to that in normal solution. Similarly, following decrease in pHo to 6.5, no recovery of intracellular acidosis was observed. We conclude that low [Cl-]o environment causes activation of extracellular HCO3- -dependent pHi-regulating mechanism, that results in the rapid recovery of pHi following acidosis, and the attenuation of acidosis-induced contraction of WKY aorta.
Despite the discrepancies observed between the data generated from humans and animals, it is a usual practice that the results obtained from animal models are extrapolated on humans. This review stresses that while animal models are essential for the research and development, a critical caution needs to be practiced in interpreting the results. Uncritical reliance on the results of animal experimentation can be dangerously misleading and has resulted in damages to human health in several cases. This review also discusses the role of certain confounding factors in using animal models due to which appropriate precautions need to be taken while deciding to conduct a study using animal models and caution is warranted in extrapolating the data obtained from pre-clinical studies on humans.
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The contractile response to acidosis in isolated aorta from spontaneously hypertensive rat (SHR) depends upon tyrosine phosphorylation of phosphatidylinositol 3 kinase (PI3-kinase) and Ca2+ influx via voltage-dependent Ca2+ channels (VDCC). In this study, verapamil, a VDCC inhibitor, was shown to markedly inhibit acidic pH-induced contraction, whereas the residual contraction in the presence of verapamil was unaffected by the PI3-kinase inhibitor, 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one hydrochloride (LY-294002). Interestingly, the LY-294002-insensitive component of contraction was further inhibited by verapamil in the presence of LY-294002. Western blotting revealed that acidosis stimulated tyrosine phosphorylation of p85, which was abolished when tissues were pretreated with tyrphostin 23, a tyrosine kinase inhibitor, verapamil or EGTA. In fura-2-loaded aortic strips, acidosis induced a rise in intracellular Ca2+ ([Ca2+]i) that was partially inhibited by LY-294002. The residual increase in [Ca2+]i caused by acidosis in the presence of LY-294002 was abolished by verapamil. These findings suggest that acidosis-induced Ca2+ influx through VDCC is the upstream event leading to the tyrosine phosphorylation of PI3-kinase, which in turn contributes to the enhancement of Ca2+ entry to some extent in SHR aorta.
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This study was performed to test the hypothesis that activation of protein kinase C (PKC) is a mechanism underlying the acidic pH-induced contraction (APIC) in spontaneously hypertensive rat (SHR) aorta. Changing pH of the bathing solution from 7.4 to 6.5 induced a marked contraction of SHR aorta. PKC inhibitors, GF109203X and calphostin C markedly inhibited the APIC selectively, without having a marked effect on the KCl-induced contraction. Inhibitors of mitogen-activated protein kinase kinase, U0126 and PD98059 mildly but significantly attenuated the APIC. However, at the similar concentrations both U0126 and PD98059 inhibited the KCl-induced contraction in a manner similar to that observed in APIC. D-609, an inhibitor of phosphatidylcholine-specific phospholipase C (PC-PLC) markedly inhibited the APIC and the extent of inhibition by this compound was similar to that shown by PKC inhibitors. Whereas, U-73122 and propranolol, inhibitors of phosphatidylinositol-specific PLC and phosphatidate phosphohydrolase, respectively, had no affect on the APIC. A tyrosine kinase inhibitor, tyrphostin 23 and GF109203X inhibited the APIC in an additive manner, and together they abolished the contractile response. From all these results, it is suggested that a significant component of the contraction observed in response to acidosis in SHR aorta is dependent upon the activation of PKC that seems to be the downstream event of the activation of PC-PLC. Furthermore, PKC- and tyrosine kinase-dependent pathways underlying the APIC are independent of each other.
The effects of acidosis were investigated on the resting and precontracted aortas from Wistar and Wistar Kyoto (WKY) rats. Decrease in pH from 7.4 to 6.5, having no effect on the resting tension of Wistar aorta, induced a marked contraction of WKY aorta. Acidic pH markedly relaxed the contraction to 300 nM phenylephrine in Wistar aorta, whereas in WKY aorta, it produced a biphasic response, an initial relaxation followed by potentiation of the contraction. In aortas loaded with fura 2-AM, phenylephrine caused an increase in intracellular Ca2+ ([Ca2+]i) and a contraction in both Wistar and WKY rats. pH 6.5 produced a decrease in [Ca2+]i to a near-basal level and almost abolished the phenylephrine-induced contraction in Wistar rat aorta. However, in WKY aorta, a biphasic response, an initial decline and later a recovery of [Ca2+]i level, was observed. Interestingly, at similar sustained [Ca2+]i, the contractile response to phenylephrine in WKY aorta was potentiated under acidic pH conditions. Acidic pH-induced inhibition of the contraction to phenylephrine was unaffected by iberiotoxin, 4-aminopyridine, and glibenclamide (Ca2+-activated, delayed rectifier and ATP-sensitive K+ channel inhibitors, respectively), in aortas from both Wistar and WKY. Decrease in extracellular pH was associated with a rapid fall in intracellular pH (pHi) and the intracellular acidification profile was not different in both strains. All these results show that acidic pH induces strain-specific inhibitory and excitatory effects on the contractile state of aortas from Wistar and WKY rats, respectively. The sustained and transient relaxant responses to acidic pH in Wistar and WKY aortas, respectively, are due to decrease in [Ca2+]i levels, but this decrease in [Ca2+]i is independent of the activation of K+ channels.
Acidic pH induces a contraction in aorta from spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats. The contractile response to acidic pH in SHR aorta is greater than that in WKY aorta. The purpose of this study was to investigate the correlation among extracellular pH (pH(o)), intracellular pH (pH(i)) and contraction in order to understand the exaggerated contractile response to acidic pH in SHR aorta. pH(i) measurement showed that at pH(o) 6.5, intracellular acidification was greater in SHR aorta than in WKY aorta. Decreasing pH(o) further to 6.2 in WKY aorta produced intracellular acidification close to that achieved at pH(o) 6.5 in SHR aorta, and at this level, the difference in contractile response between the two strains was also abolished. These results suggest that acidic pH(i), but not pH(o), is closely correlated with the contractile response and that the exaggerated contractile response in SHR aorta is due to a greater fall in pH(i).
Acidic pH induced a contraction in the isolated aorta from Wistar Kyoto rat. The magnitude of contraction was dependent upon the degree of extracellular acidification. The maximum level of contraction observed at pH 6.5 was 84.6 +/- 3.4% of the 64.8 mM KCl-induced contraction. To investigate the role of extracellular as well as intracellular Ca(2+) in acidic pH-induced contraction (APIC), we changed the extracellular pH in the presence of EGTA. Sustained contraction induced by acidic pH in the presence of extracellular Ca(2+) was completely abolished in the presence of EGTA, while a transient but significant contraction was still observed. Ryanodine, a selective ryanodine receptor blocker and cyclopiazonic acid (CPA), an inhibitor of sarco-/endoplasmic reticulum Ca(2+) ATPase, abolished the transient contraction, when pH was decreased in Ca(2+)-free solution. On the other hand, neither xestospongin C, a selective inositol-1,4,5-trisphosphate receptor antagonist nor U-73122, a phospholipase C inhibitor showed this effect. These results suggest the involvement of Ca(2+) release from ryanodine-/CPA-sensitive store of sarcoplasmic reticulum (SR). In normal Ca(2+)-containing solution, ryanodine and CPA did not alter the maximum level of APIC. However, they significantly decreased the rate of rise of APIC. U-73122, suppressed the maximum contraction induced by acidic pH without affecting the rate of rise of APIC, while xestospongin C and U-73343, an inactive analogue of U-73122, had no effect on both parameters of APIC. From these results, it is concluded that acidic pH induces Ca(2+) release from the ryanodine-/CPA-sensitive store of SR and that release provides supportive effect on initiating rapid transient contraction, but not on the sustained contraction, which is entirely due to Ca(2+) influx.
Hydrogen ion concentration (H+) is an important regulator of many cellular functions including contractility of the vascular smooth muscle cells. In many physiological and pathological states, changes in pH may have profound effects on the contractility of blood vessels and thus on the peripheral vascular resistance and blood pressure. Therefore, it is necessary to understand the likely outcome of the situations in the conditions, where pH changes are frequent. In this review, the mechanisms involved in the contractile responses of vascular tissues to pH changes are discussed.
V-1 positively controls catecholamine synthetic gene transcription to promote catecholamine production in PC12D cells. In this study, immunohistochemical analysis revealed that in Wistar rats, V-1 immunoreactivity was localized not only in sympathetic axons but also in the cytoplasm of cardiomyocytes, and that the immunoreactivity in atrial myocytes was more intense than that in ventricular myocytes. Western blot analysis also showed that V-1 expression level in the atrium was higher than that in the ventricle of Wistar rat hearts. When Dahl salt-sensitive (DS) rats were fed an 8% NaCl diet after the age of 6 weeks, blood pressure was raised 230mm Hg at 18 weeks. V-1 expression was shown to be increased in the atrial myocytes of these DS rats, but not in the sympathetic axons, when assayed by immunohistochemistry. These results suggest that in normotensive rats, V-1 is preferentially expressed in the cytoplasm of cardiomyocytes in the atrium rather than in the ventricle. It is also suggested that V-1 expression is increased by hypertension in DS rat atrium.
pH regulates various cellular functions. Previously, we have described that acidic pH produces depolarization and contraction in isolated aorta from spontaneously hypertensive (SHR) and Wistar Kyoto (WKY) rats [Br. J. Pharmacol. 118 (1996) 485]. The aim of the present study was to investigate the involvement of Cl- channels in acidic pH-induced contraction. Changing the pH of the bathing solution from 7.4 to 6.5 induced a contraction in both SHR and WKY aorta, which was 127.50+/-13.32% and 79.27+/-0.94% of the 64.8 mM KCl-induced contraction, respectively. The acidic pH-induced contraction was partially inhibited by the voltage-dependent Ca2+ channel (VDCC) blockers, verapamil (1 microM) and nifedipine (0.1 microM). The Cl- channel inhibitors, diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS) (0.5 mM), 9-anthracene chloride (0.5 mM), indanyloxyacetic acid (30 microM) and niflumic acid (3 microM) also inhibited the acidic pH-induced contraction and the degree of attenuation was comparable to that of VDCC blockers. DIDS, 9-anthracene chloride and niflumic acid at concentrations used to inhibit the acidic pH-induced contraction also inhibited the 10 microM phenylephrine-induced contraction partially, without affecting the 64.8 mM KCl-induced contraction, whereas both the contractions were inhibited by indanyloxyacetic acid with equal efficacy. Indanyloxyacetic acid but not DIDS, 9-anthracene chloride or niflumic acid inhibited the 24.8 mM KCl-induced contraction. Simultaneous measurement of cytosolic Ca2+ and tension showed that niflumic acid reversed the increase in intracellular Ca2+ level and inhibited the contraction caused by acidic pH. Similarly, acidic pH depolarized the cultured vascular smooth muscle cells from SHR and the depolarization was completely reversible after the administration of niflumic acid. All these results suggest that the activation of Cl- channels is an important mechanism underlying the depolarization and contraction induced by acidic pH in SHR and WKY aortas.
Acidic pH induced a contraction (APIC) in isolated aortas from spontaneously hypertensive (SHR) and Wistar Kyoto rats, but failed to produce any response in age-matched Wistar rat aorta. This study was conducted to test the hypothesis that tyrosine phosphorylation of proteins is a molecular mechanism underlying the APIC. Tyrosine kinase inhibitors, genistein and tyrphostin 23 inhibited the APIC in a concentration-dependent manner. APIC was inhibited by phosphatidylinositol 3-kinase (PI3-kinase) inhibitors, LY-294002 [2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one hydrochloride] and wortmannin. Consistent with the results from tension measurement experiments, Western blot analysis showed that acidic pH induced an appreciable increment of tyrosine phosphorylation of 85-kDa protein (p85) in SHR aorta, which was completely inhibited by tyrphostin 23, whereas in Wistar rat aorta, the protein tyrosine phosphorylation was not observed. Further investigations using immunoprecipitation followed by Western blotting confirmed an increase in the tyrosine phosphorylation of p85. Analysis by SDS-polyacrylamide gel electrophoresis followed by silver staining of the gel revealed that amounts of multiple proteins with molecular sizes of 120, 130, 210, and 225 kDa were increased at acidic pH, which were immunoprecipitated with anti-phosphotyrosine antibody. Western blotting using a specific anti-PI3-kinase antibody identified the p85 as the regulatory subunit of PI3-kinase, whereas 120-, 130-, and 225-kDa proteins were identified by mass spectrometry as pro-alpha2 (I) collagen, collagen alpha1 (I) chain, and fibernectin I, respectively. As assayed by Western blotting using anti-myosin light chain (MLC) antibody, acidic pH induced a stimulation of MLC phosphorylation, and the stimulated MLC phosphorylation was abolished by tyrphostin 23 and LY-294002. These results suggest that acidic pH induces an increase in tyrosine phosphorylation of PI3-kinase, resulting in the MLC phosphorylation-dependent contraction of SHR aorta.