The water channel AQP1 is expressed in human atherosclerotic vascular lesions and AQP1 deficiency augments angiotensin II-induced atherosclerosis in mice.

AIM: The water channel aquaporin 1 (AQP1) promotes endothelial cell migration. It was hypothesized that AQP1 promotes neovascularization and growth of atherosclerotic plaques. METHODS: AQP1 immunoreactivity and protein abundance was examined in human and murine atherosclerotic lesions and aortic aneurysms. Apolipoprotein E (ApoE) knockout (-/-) and AQP1-/-ApoE-/- mice were developed and fed Western diet (WD) for 8 and 16 weeks to accelerate the atherosclerosis process. In ApoE-/- and AQP1-/-ApoE-/- mice abdominal aortic aneurysms (AAA) were induced by angiotensin II (ANGII) infusion by osmotic minipumps for 4 weeks. RESULTS: In human atherosclerotic lesions and AAA, AQP1 immunoreactive protein was associated with intralesional small vessels. In ApoE-/- mouse aorta, APQ1 mRNA levels were increased with time on WD (n = 7-9, P < 0.003). Both in murine lesions at the aortic root and in the abdominal aortic aneurysmal wall, AQP1 immunoreactivity was associated with microvascular structures. The atherosclerotic lesion burden was enhanced significantly in ANGII-infused AQP1-/-ApoE-/- mice compared with ApoE-/- mice, but neither incidence nor progression of AAA was different. The aortic lesion burden increased with time on WD but was not different between ApoE-/- and AQP1-/-ApoE-/- mice at either 8 or 16 weeks (n = 13-15). Baseline blood pressure and ANGII-induced hypertension were not different between genotypes. CONCLUSION: AQP1 is expressed in atherosclerotic lesion neovasculature in human and mouse arteries and AQP1 deficiency augments lesion development in ANGII-promoted atherosclerosis in mice. Normal function of AQP1 affords cardiovascular protection.

Biosensor cell assay for measuring real-time aldosterone-induced release of histamine from mesenteric arteries.

AIMS: The aims were to develop a method for real-time detection of histamine release and to test whether incubation with aldosterone induces histamine release from isolated, perfused mice mesenteric arteries. METHODS: Fura-2-loaded HEK-293 cells transfected with the histamine H1 receptor was used as a sensitive biosensor assay for histamine release from isolated mouse mesenteric arteries. Activation of the H1 receptor by histamine was measured as an increased number of intracellular Ca2+ transient peaks using fluorescence imaging. RESULTS: The developed biosensor was sensitive to histamine in physiological relevant concentrations and responded to substances released by the artery preparation. Aldosterone treatment of mesenteric arteries from wild-type mice for 50 min resulted in an increased number of intracellular Ca2+ transient peaks in the biosensor cells, which was significantly inhibited by the histamine H1 blocker pyrilamine. Mesenteric arteries from mast cell-deficient SASH mice induced similar pyrilamine-sensitive Ca2+ transient response in the biosensor cells. Mesenteric arteries from wild-type and SASH mice expressed histamine decarboxylase mRNA, indicating that mast cells are not the only source of histamine release. CONCLUSION: The developed biosensor assay can measure release of substances from vascular preparations. Histamine is released from the vessel preparation in response to aldosterone treatment independently of mast cells. The assay enables us to study a new signaling mechanism for vascular responses induced by aldosterone.

P/Q-type and T-type voltage-gated calcium channels are involved in the contraction of mammary and brain blood vessels from hypertensive patients.

AIM: Calcium channel blockers are widely used in cardiovascular diseases. Besides L-type channels, T- and P/Q-type calcium channels are involved in the contraction of human renal blood vessels. It was hypothesized that T- and P/Q-type channels are involved in the contraction of human brain and mammary blood vessels. METHODS: Internal mammary arteries from bypass surgery patients and cerebral arterioles from patients with brain tumours with and without hypertension were tested in a myograph and perfusion set-up. PCR and immunohistochemistry were performed on isolated blood vessels. RESULTS: The P/Q-type antagonist ω-agatoxin IVA (10-8  mol L-1 ) and the T-type calcium blocker mibefradil (10-7  mol L-1 ) inhibited KCl depolarization-induced contraction in mammary arteries from hypertensive patients with no effect on blood vessels from normotensive patients. ω-Agatoxin IVA decreased contraction in cerebral arterioles from hypertensive patients. L-type blocker nifedipine abolished the contraction in mammary arteries. PCR analysis showed expression of P/Q-type (Cav 2.1), T-type (Cav 3.1 and Cav 3.2) and L-type (Cav 1.2) calcium channels in mammary and cerebral arteries. Immunohistochemical labelling of mammary and cerebral arteries revealed the presence of Cav 2.1 in endothelial and smooth muscle cells. Cav 3.1 was also detected in mammary arteries. CONCLUSION: P/Q- and T-type Cav are present in human internal mammary arteries and in cerebral penetrating arterioles. P/Q- and T-type calcium channels are involved in the contraction of mammary arteries from hypertensive patients but not from normotensive patients. Furthermore, in cerebral arterioles P/Q-type channels importance was restricted to hypertensive patients might lead to that T- and P/Q-type channels could be a new target in hypertensive patients.

Myogenic tone is impaired at low arterial pressure in mice deficient in the low-voltage-activated CaV 3.1 T-type Ca(2+) channel.

AIM: Using mice deficient in the CaV 3.1 T-type Ca(2+) channel, the aim of the present study was to elucidate the molecular identity of non-L-type channels involved in vascular tone regulation in mesenteric arteries and arterioles. METHODS: We used immunofluorescence microscopy to localize CaV 3.1 channels, patch clamp electrophysiology to test the effects of a putative T-type channel blocker NNC 55-0396 on whole-cell Ca(2+) currents, pressure myography and Ca(2+) imaging to test diameter and Ca(2+) responses of the applied vasoconstrictors, and Q-PCR to check mRNA expression levels of several Ca(2+) handling proteins in wild-type and CaV 3.1(-/-) mice. RESULTS: Our data indicated that CaV 3.1 channels are important for the maintenance of myogenic tone at low pressures (40-80 mm Hg), whereas they are not involved in high-voltage-activated Ca(2+) currents, Ca(2+) entry or vasoconstriction to high KCl in mesenteric arteries and arterioles. Furthermore, we show that NNC 55-0396 is not a specific T-type channel inhibitor, as it potently blocks L-type and non-L-type high-voltage-activated Ca(2+) currents in mouse mesenteric vascular smooth muscle cell. CONCLUSION: Our data using mice deficient in the CaV 3.1 T-type channel represent new evidence for the involvement of non-L-type channels in arteriolar tone regulation. We showed that CaV 3.1 channels are important for the myogenic tone at low arterial pressure, which is potentially relevant under resting conditions in vivo. Moreover, CaV 3.1 channels are not involved in Ca(2+) entry and vasoconstriction to large depolarization with, for example, high KCl. Finally, we caution against using NNC 55-0396 as a specific T-type channel blocker in native cells expressing high-voltage-activated Ca(2+) channels.

Functional and pharmacological consequences of the distribution of voltage-gated calcium channels in the renal blood vessels.

Calcium channel blockers are widely used to treat hypertension because they inhibit voltage-gated calcium channels that mediate transmembrane calcium influx in, for example, vascular smooth muscle and cardiomyocytes. The calcium channel family consists of several subfamilies, of which the L-type is usually associated with vascular contractility. However, the L-, T- and P-/Q-types of calcium channels are present in the renal vasculature and are differentially involved in controlling vascular contractility, thereby contributing to regulation of kidney function and blood pressure. In the preglomerular vascular bed, all the three channel families are present. However, the T-type channel is the only channel in cortical efferent arterioles which is in contrast to the juxtamedullary efferent arteriole, and that leads to diverse functional effects of L- and T-type channel inhibition. Furthermore, by different mechanisms, T-type channels may contribute to both constriction and dilation of the arterioles. Finally, P-/Q-type channels are involved in the regulation of human intrarenal arterial contractility. The calcium blockers used in the clinic affect not only L-type but also P-/Q- and T-type channels. Therefore, the distinct effect obtained by inhibiting a given subtype or set of channels under experimental settings should be considered when choosing a calcium blocker for treatment. T-type channels seem to be crucial for regulating the GFR and the filtration fraction. Use of blockers is expected to lead to preferential efferent vasodilation, reduction of glomerular pressure and proteinuria. Therefore, renovascular T-type channels might provide novel therapeutic targets, and may have superior renoprotective effects compared to conventional calcium blockers.

Adenosine elicits an eNOS-independent reduction in arterial blood pressure in conscious mice that involves adenosine A2A receptors.

AIMS: Adenosine plays an important role in the regulation of heart rate (HR) and vascular reactivity. However, the mechanisms underlying the acute effect of adenosine on arterial blood pressure in conscious mice are unclear. Therefore, this study investigated the effect of the nucleoside on mean arterial blood pressure (MAP) and HR in conscious mice. METHODS: Chronic indwelling catheters were placed in C57Bl/6J (WT) and endothelial nitric oxide synthase knockout (eNOS(-/-)) mice for continuous measurements of MAP and HR. Using PCR and myograph analysis, involvement of adenosine receptors was investigated in human and mouse renal blood vessels. RESULTS: Bolus infusion of 0.5 mg kg(-1) adenosine elicited significant transient decreases in MAP (99.3 ± 2.3 to 70.4 ± 4.5 mmHg) and HR (603.2 ± 18.3 to 364.3 ± 49.2 min(-1)), which were inhibited by the A(2A) receptor antagonist ZM 241385. Activation of adenosine A(2A) receptors with CGS 21680 (0.02 mg kg(-1)) caused a significant reduction in MAP from 99.6 ± 1.2 to 73.1 ± 3.6 mmHg accompanied by tachycardia (610.5 ± 9.3 to 677.5 ± 9.5 min(-1)). The reduction in MAP observed after adenosine or CGS 21680 administrations was not significantly different in WT and eNOS(-/-) mice. In isolated human and mouse intrarenal arteries, adenosine caused a relaxation dependent on A(2A) adenosine receptor activation. A(2A) receptors were present in both human and mouse arteries whereas A(1) and A(2B) receptors were only present in mouse arteries. CONCLUSION: In conclusion, acute adenosine administration and selective stimulation of adenosine A(2A) receptors results in an immediate, transient eNOS-independent reduction in MAP. A(2A) receptor activation causes relaxation of human and mouse arteries.

Uridine adenosine tetraphosphate affects contractility of mouse aorta and decreases blood pressure in conscious rats and mice.

AIM: in the anaesthetized rat, uridine adenosine tetraphosphate (Up(4) A) is a circulating, endothelium-derived vasoconstrictor presumably operating as such in un-anaesthetized animals. The present study investigated the in vivo effects of Up(4) A in conscious mice and rats, and its direct vascular effects in the mouse aorta in vitro. METHODS: in vivo, Up(4) A was given as step-up infusion at rates of 8-512 nmol min(-1) kg(-1) for 30 min periods in chronically catheterized rodents. In vitro, the effect of Up(4) A on rings of mouse aortae mounted in a myograph was tested. RESULTS: high doses of Up(4) A (mice: 512 nmol min(-1) kg(-1) ; rats: 128 nmol min(-1) kg(-1) ) caused hypotension (99 (+/-)4 to 64 7(+/-) mmHg and 114 (+/-) 3 to 108 (+/-) 3 mmHg, respectively, both P < 0.01). In rats, Up(4) A significantly decreased sodium excretion by >75% and potassium excretion by approximately 60% without significant changes in urine flow. Exposure of phenylephrine-contracted rings to increasing concentrations of Up(4) A elicited contraction at 10(-7) and 10(-6) molL(-1) (18 ± 2% and 76 (+/-) 16% respectively); unexpectedly, 10(-5) molL(-1) caused a biphasic response with a contraction (19 6(+/-)2%) followed by a relaxation (-46 (+/-) 6%). No relaxation was observed when the concentration was increased further. Bolus exposure to 10(-5) molL(-1) of Up(4) A caused contraction (+80 (+/-) 2%). Added successively to untreated vessels, increasing concentrations of Up(4) A (10(-7) -10(-5) molL(-1) ) induced a biphasic response of contraction followed by relaxation. CONCLUSION: up(4) A has direct biphasic effects on vascular smooth muscle of the mouse aorta but vasoconstriction dominates at low concentrations. In conscious rodents, step-up infusions of Up(4) A elicit hypotension and electrolyte retention.

Intracellular signalling pathways in the vasoconstrictor response of mouse afferent arterioles to adenosine.

AIMS: Adenosine causes vasoconstriction of afferent arterioles of the mouse kidney through activation of adenosine A(1) receptors and Gi-mediated stimulation of phospholipase C. In the present study, we further explored the signalling pathways by which adenosine causes arteriolar vasoconstriction. METHODS AND RESULTS: Adenosine (10(-7) M) significantly increased the intracellular calcium concentration in mouse isolated afferent arterioles measured by fura-2 fluorescence. Pre-treatment with thapsigargin (2 microM) blocked the vasoconstrictor action of adenosine (10(-7) M) indicating that release of calcium from the sarcoplasmic reticulum (SR), stimulated presumably by IP(3), is involved in the adenosine contraction mechanism of the afferent arteriole. In agreement with this notion is the observation that 2 aminoethoxydiphenyl borate (100 microM) blocked the adenosine-induced constriction whereas the protein kinase C inhibitor calphostin C had no effect. The calcium-activated chloride channel inhibitor IAA-94 (30 microM) inhibited the adenosine-mediated constriction. Patch clamp experiments showed that adenosine treatment induced a depolarizing current in preglomerular smooth muscle cells which was abolished by IAA-94. Furthermore, the vasoconstriction caused by adenosine was significantly inhibited by 5 microM nifedipine (control 8.3 +/- 0.2 microM, ado 3.6 +/- 0.6 microM, ado + nifedipine 6.8 +/- 0.2 microM) suggesting involvement of voltage-dependent calcium channels. CONCLUSION: We conclude that adenosine mediates vasoconstriction of afferent arterioles through an increase in intracellular calcium concentration resulting from release of calcium from the SR followed by activation of Ca(2+)-activated chloride channels leading to depolarization and influx of calcium through voltage-dependent calcium channels.

Vasoconstrictor and vasodilator effects of adenosine in the mouse kidney due to preferential activation of A1 or A2 adenosine receptors.

The present experiments in mice were performed to determine the steady-state effects of exogenous adenosine on the vascular resistance of the whole kidney, of superficial blood vessels, and of afferent arterioles. The steady-state effect of an intravenous infusion of adenosine (5, 10, and 20 microg/min) in wild-type mice was vasodilatation as evidenced by significant reductions of renal and superficial vascular resistance. Resistance decreases were augmented in adenosine 1 receptor (A1AR) -/- mice. Renal vasodilatation by the A2aAR agonist CGS 21680A [2-p-(2-carboxyethyl)phenethyl-amino-5'-N-ethylcarboxamido-adenosine hydrochloride] (0.25, 0.5, and 1 microg/kg/min) and inhibition of adenosine-induced relaxation by the A2aAR antagonist ZM-241385 [4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-yl-amino]ethyl)phenol] (20 mg/kg) suggests that the reduction of renovascular resistance was largely mediated by A2aAR. After treatment with Nomega-nitro-L-arginine methyl ester (L-NAME) adenosine was unable to alter superficial blood flow and resistance significantly indicating that adenosine-induced dilatation is NO-dependent. Absence of a dilatory effect in endothelial nitric-oxide synthase (NOS) -/- mice suggests endothelial NOS as the source of NO. When infused into the subcapsular interstitium, adenosine reduced superficial blood flow through A1AR activation. Adenosine (10(-7) M) constricted isolated perfused afferent arterioles when added to the bath but not when added to the luminal perfusate. Luminal adenosine caused vasoconstriction in the presence of L-NAME or the A2AR antagonist 3,7-dimethyl-1-(2-propynyl)xanthine. Our data show that global elevation of renal adenosine causes steady-state vasorelaxation resulting from adenosine 2 receptor (A2AR)-mediated generation of NO. In contrast, selective augmentation of adenosine around afferent arterioles causes persistent vasoconstriction, indicating A1AR dominance. Thus, adenosine is a renal constrictor only when it can interact with afferent arteriolar A1AR without affecting the bulk of renal A2AR at the same time.

Anagrelide treatment in 52 patients with chronic myeloproliferative diseases.

In this retrospective multi-centre study, we report our experience with anagrelide in the treatment of thrombocytosis in patients with chronic myeloproliferative diseases. Our study included 52 patients (age 20-78 years). The initial anagrelide dose was, in general, 0.5 mg once daily and mean maintenance dosage was 1.7 mg/day. The overall response rate was 79% including 75% complete remission and 4% partial remission. Forty-two patients (81%) had adverse effects and in 29% of the study population, the adverse effects necessitated cessation of anagrelide. The most common adverse effect was moderate anaemia (50%). Two patients experienced erectile dysfunction which has been described only once previously in association with anagrelide treatment. One patient progressed to acute leukaemia. However, this patient had been pre-treated with two potentially leukaemogenic drugs and had only been in short-term treatment with anagrelide. Furthermore, a total of 13 events were recorded. More than 25% of these events occurred in patients with platelet counts between 400 and 600 x 10(9)/l and almost 40% of all events occurred in patients with platelet counts above 400 x 10(9)/l. This observation supports the hypothesis that aggressive control of thrombocytosis to a platelet count <400 x 10(9)/l might reduce the number of thrombohaemorrhagic events. Anagrelide is safe and effective in reducing the platelet counts, but a high proportion of the patients discontinue treatment because of the adverse effects of the drug.

Rapid non-genomic effects of aldosterone on rodent vascular function.

The main role of aldosterone is to maintain body sodium homeostasis by promoting salt reabsorption in the collecting ducts of the kidney. In the cardiovascular system, aldosterone may be harmful in a number of disease states by inducing fibrosis and vascular dysfunction. The present review describes novel results from several laboratories, which show that aldosterone also has beneficial effects in the cardiovascular system by stimulating the production of nitric oxide (NO) from the endothelium. The effect of aldosterone is seen within minutes, and is not inhibited by blockers of gene transcription, thus pointing to a non-genomic mechanism. Furthermore, this potentially beneficial effect is observed at low physiological concentrations of aldosterone (0.1-10 pm). The effect is mediated by the classical mineralocorticoid receptor, and it involves heat shock protein 90, phosphatidylinositol (PI)-3 kinase, protein kinase B, endothelial nitric oxide synthase, and liberation of NO. It is proposed that in healthy individuals with a functioning NO system, the detrimental effects of aldosterone on cardiovascular function are balanced by activation of the potentially beneficial effect of NO. However, in situations with endothelial dysfunction, such as congestive heart failure and hypertension, the negative effects of aldosterone are unopposed and inhibition of aldosterone is warranted.

Plasma renin in mice with one or two renin genes.

AIM: In the present study we have investigated whether the presence of a second renin gene exerts an overriding influence on plasma renin such that mice with two renin genes have consistently higher renin levels than mice with only one renin gene. METHODS: Plasma renin was determined as the rate of angiotensin I generation using a radioimmunoassay (RIA) kit with (plasma renin concentration, PRC) or without (plasma renin activity, PRA) the addition of purified rat angiotensinogen as substrate. RESULTS: In male 129SvJ, DBA/2 and Swiss Webster mice, strains possessing both Ren-1 and Ren-2, PRC (ng Ang I mL(-1) h(-1)) averaged 178 +/- 36, 563 +/- 57 and 550 +/- 43 while PRA was 2.9 +/- 0.5, 3.6 +/- 0.8 and 7.8 +/- 1.2. In male C57BL/6, C3H and BALB/c mice that express only Ren-1, PRC averaged 426 +/- 133, 917 +/- 105 and 315 +/- 72, and PRA was 3.4 +/- 1.0, 6.9 +/- 1.7 and 4.5 +/- 1.2. In the two renin gene A1AR-/- mice compared with the one renin gene A1AR+/+, PRC averaged 538 +/- 321 and 415 +/- 159 while PRA averaged 3.2 +/- 1.1 and 4.4 +/- 1.4 ng Ang I mL(-1) h(-1). Aldosterone levels showed no significant differences between one renin (C57BL/6, C3H and BALB/c) and two renin (129SvJ, DBA/2 and Swiss Webster) gene mice. Furthermore, by quantitative real-time polymerase chain reaction (RT-PCR) we found no correlation between the number of renin genes and whole kidney renin mRNA levels from one and two renin gene mice. CONCLUSION: Our data show that baseline plasma renin is not systematically higher in mice with two renin genes than in one renin gene mice. Thus, the presence of a second renin gene does not seem to be a major determinant of differences in PRC between different mouse strains.

Rapid inhibition of vasoconstriction in renal afferent arterioles by aldosterone.

Aldosterone has been suggested to elicit vessel contraction via a nongenomic mechanism. We tested this proposal in microdissected, perfused rabbit renal afferent arterioles. Aldosterone had no effect on internal diameter in concentrations from 10(-10) to 10(-5) mol/L, but aldosterone abolished the ability of 100 mmol/L KCl to induce vascular contraction. The inhibitory effect of aldosterone was observed from 1 pmol/L. The inhibitory effect was significant after 5 minutes and maximal after 20 minutes and was fully reversible. Actinomycin D (10(-6) mol/L) prolonged the effect of aldosterone. The effect was abolished by the mineralocorticoid receptor antagonist spironolactone (10(-7) mol/L) but not by the glucocorticoid receptor antagonist mifepristone (10(-6) mol/L). The K+-mediated increase of intracellular calcium concentration in afferent arterioles was not affected by aldosterone. Mineralocorticoid receptor was detected by reverse transcription-polymerase chain reaction and immunohistochemistry in rat renal vasculature and rabbit endothelial cells. Inhibition of phosphatidylinositol (PI)-3 kinase with LY 294002 (3x10(-6) mol/L) restored sensitivity to K+ in the presence of aldosterone, and afferent arterioles were immunopositive for PI-3 kinase subunit p110alpha. Inhibition of NO formation by L-NAME (10(-4) mol/L) or inhibition of soluble guanylyl cyclase with 1H-(1,2,4)Oxadiazolo[4,3-a]quinoxaline-1-one restored K+-induced vasoreactivity in the presence of aldosterone. Similar to aldosterone, the NO donor sodium nitroprusside inhibited K+-induced vascular contraction. Geldanamycin (10(-6) mol/L), an inhibitor of heat shock protein 90, abolished aldosterone-induced vasorelaxation. We conclude that aldosterone inhibits depolarization-induced vasoconstriction in renal afferent arterioles by a rapid nongenomic mechanism that is initiated by mineralocorticoid receptor activation and involves PI-3 kinase, protein kinase B, and heat shock protein 90-mediated stimulation of NO generation.

Differential expression of T- and L-type voltage-dependent calcium channels in renal resistance vessels.

The distribution of voltage-dependent calcium channels in kidney pre- and postglomerular resistance vessels was determined at the molecular and functional levels. Reverse transcription-polymerase chain reaction analysis of microdissected rat preglomerular vessels and cultured smooth muscle cells showed coexpression of mRNAs for T-type subunits (Ca(V)3.1, Ca(V)3.2) and for an L-type subunit (Ca(V)1.2). The same expression pattern was observed in juxtamedullary efferent arterioles and outer medullary vasa recta. No calcium channel messages were detected in cortical efferent arterioles. Ca(V)1.2 protein was demonstrated by immunochemical labeling of rat preglomerular vasculature and juxtamedullary efferent arterioles and vasa recta. Cortical efferent arterioles were not immunopositive. Recordings of intracellular calcium concentration with digital fluorescence imaging microscopy showed a significant increase of calcium in response to K(+) (100 mmol/L) in isolated afferent arterioles (140+/-25%) and in juxtamedullary efferent arterioles (118+/-21%). These calcium responses were attenuated by the L-type antagonist calciseptine and by the T-type antagonist mibefradil. Intracellular calcium increased in response to K(+) in cortical efferent arterioles (21+/-9%). Mibefradil and nickel concentration dependently blocked K(+)-induced contraction of perfused rabbit afferent arterioles. Calciseptine blocked the contraction mediated by K(+) (EC(50) 8x10(-14)). S-(-)-Bay K 8644 had no effect on vascular diameter in the afferent arteriole. We conclude that voltage-dependent L- and T-type calcium channels are expressed and of functional significance in renal cortical preglomerular vessels, in juxtamedullary efferent arterioles, and in outer medullary vasa recta, but not in cortical efferent arterioles.

[Clinical use of recombinant human thrombopoietin. Status and perspectives]. / Klinisk anvendelse af rekombinant humant trombopoietin. Status og perspektiver.

Thrombopoietin (TPO) is primarily produced by hepatocytes and regulates the production and differentiation of megakaryocytes and platelets in the bone marrow. The endogenous TPO level is increased when the megakaryocyte count is low, and high in aplastic anaemia and after myeloablative chemotherapy. TPO is cloned and manufactured by a recombinant technique for clinical use. Treatment with recombinant human TPO (rhTPO) after intensive chemotherapy may reduce the need for platelet transfusions. Administration of granulocyte colony-stimulating factor in combination with rhTPO has enhanced the mobilisation and harvest product of haematopoietic stem cells. Whether rhTPO is effective in the treatment of the myelodysplastic syndrome, aplastic anaemia, and other conditions with bone marrow insufficiency (including AIDS) is not yet known. In liver cirrhosis, the endogenous TPO level rapidly increases after liver transplantation. Accordingly, substitution of rhTPO may be indicated in advanced liver failure complicated by thrombocytopenia and bleeding.

Localization of prostaglandin E(2) EP2 and EP4 receptors in the rat kidney.

We investigated the localization of cAMP-coupled prostaglandin E(2) EP2 and EP4 receptor expression in the rat kidney. EP2 mRNA was restricted to the outer and inner medulla in rat kidney, as determined by RNase protection assay. RT-PCR analysis of microdissected resistance vessels and nephron segments showed EP2 expression in descending thin limb of Henle's loop (DTL) and in vasa recta of the outer medulla. The EP4 receptor was expressed in distal convoluted tubule (DCT) and cortical collecting duct (CCD) in preglomerular vessels, and in outer medullary vasa recta. Butaprost, an EP2 receptor-selective agonist, dose dependently raised cAMP levels in microdissected DTL and outer medullary vasa recta specimens but had no effect in EP2-negative outer medullary collecting duct segments. Dietary salt intake did not alter EP2 expression in the kidney medulla. These results suggest that PGE(2) may act in the resistance vessels and in the DTL and DCT-CCD segments as a paracrine, cAMP-dependent regulator of vascular resistance and tubular transport, respectively.