充血性心力衰竭患者尿液水通道蛋白-2的临床检测

目的 :探讨充血性心力衰竭 （ CHF）患者尿液水通道蛋白 -2 （ aquaporin-2 ,AQP2 ）浓度的变化及其与低钠血症的关系。方法 :应用双抗体夹心 ELISA法检测 CHF患者 （ 3 64例 ）和健康对照者 （ 82例 ）的尿液 AQP2浓度 ,并测定血钠、左室射血分数 （ L VEF）等指标。结果 :CHF患者比对照组尿 AQP2的浓度明显增高 （ P<0 .0 1） ,且 CHF越重（ NYHY等级越高、L VEF越小 ）及血钠越低 ,尿 AQP2的浓度越高。结论 :CHF患者水潴留及低钠血症越重 ,尿液AQP2排出越多。监测尿液 AQP2浓度 ,为临床防治 CHF及水钠失衡提供了新的检测指标和方法。     

Urinary excretion of aquaporin-2 water channel differentiates psychogenic polydipsia from central diabetes insipidus.

The present study was undertaken to determine whether urinary excretion of aquaporin-2 (AQP-2) water channel under ad libitum water intake is of value to differentiate polyuria caused by psychogenic polydipsia from central diabetes insipidus. A 30-min urine collection was made at 0900 h in 3 groups of: 11 patients with central diabetes insipidus (22-68 yr old), 10 patients with psychogenic polydipsia (28-60 yr old), and 15 normal subjects (21-38 yr old). In the patients with central diabetes insipidus, the plasma arginine vasopressin level was low despite hyperosmolality, resulting in hypotonic urine. Urinary excretion of AQP-2 was 37 +/- 15 fmol/mg creatinine, a value one-fifth less than that in the normal subjects. In the patients with psychogenic polydipsia, plasma arginine vasopressin and urinary osmolality were as low as those in the patients with central diabetes insipidus. However, urinary excretion of AQP-2 of 187 +/- 45 fmol/mg creatinine was not decreased, and its excretion was equal to that in the normal subjects. These results indicate that urinary excretion of AQP-2, under ad libitum water drinking, participates in the differentiation of psychogenic polydipsia from central diabetes insipidus.     

Urinary aquaporin-2 excretion in nocturnal enuresis

OBJECTIVE: To evaluate the role of the arginine vasopressin (AVP)-aquaporin-2 (AQP-2) axis in the pathogenesis of nocturnal enuresis. STUDY PARTICIPANTS: Twelve children (seven male and five female), aged 11.6+/-4.3 (6.7-15.6) years, suffering from primary monosymptomatic nocturnal enuresis and 12 healthy children, matched for sex and age. Enuretic children were further subdivided into responders and non-responders to treatment with 1-desamino-8-d-AVP (DDAVP). METHODS: Serum concentrations of AVP, and plasma and urine osmolality were measured at night (0100, 0400 and 0700 h), together with nocturnal urinary excretion of AQP-2 (2000-0800 h). Magnetic resonance imaging (MRI) of the pituitary gland was carried out to evaluate the amount of AVP stored in the posthypophysis. RESULTS: Mean AVP serum concentrations were similar in patients and controls. Urinary AQP-2 was also similar in patients and controls, but responders had a significantly lower level of AQP-2 than non-responders (P<0.005). Plasma osmolality was greater in patients than in controls (P<0.001), whereas urinary osmolality was similar in both groups. No difference in the ratio of the signal intensity of the posterior lobe of the hypophysis to that of the pons (AVP content) was found between patients and controls or between responders and non-responders. CONCLUSION: A decreased urinary excretion of AQP-2 is associated with, and seems to have a role in, nocturnal enuresis, at least in some children, and this could also explain why only some of them respond to DDAVP treatment.     

间接酶联免疫吸附法定量检测大鼠尿液水通道蛋白-2浓度

目的通过间接酶联免疫吸附测定(ELISA)法,检测正常大鼠不同水代谢状态下尿液水通道蛋白-2(AQP2)。方法收集大鼠正常,禁水24 h以及水负荷状态(80 ml/kg)下的尿液,用间接ELISA法检测尿液中AQP2。结果本方法批内差异系数6.5%,批间差异系数7%,灵敏度为112.5 pmol/L;禁水24 h后大鼠尿液AQP2/肌酐显著增加(P<0.01),给予水负荷后尿液AQP2/肌酐显著降低(P<0.01);且尿渗量与尿液AQP2/肌酐呈正相关(r=0.952,P<0.001)。结论间接ELISA法可以较好地检测大鼠尿液中AQP2。     

问接酶联免疫吸附法定量检测大鼠尿液水通道蛋白-2浓度

目的通过间接酶联免疫吸附测定（ELISA）法，检测正常大鼠不同水代谢状态下尿液水通道蛋白-2（AQP2）。方法收集大鼠正常，禁水24h以及水负荷状态（80ml／kg）下的尿液，用间接ELISA法检测尿液中AQP2。结果本方法批内差异系数6．5％，批间差异系数7％，灵敏度为112．5pmol／L；禁水24h后大鼠尿液AQP2／肌酐显著增加（P〈0．01），给予水负荷后尿液AQP2／肌酐显著降低（P〈0．01）；且尿渗量与尿液AQP2／肌酐呈正相关（r=0．952，P〈0．001）。结论间接ELISA法可以较好地检测大鼠尿液中AQP2。     

慢性心力衰竭大鼠模型尿液水通道蛋白_2的检测

目的　探讨慢性心力衰竭大鼠模型尿液水通道蛋白2 (aquaporin 2 )浓度的改变及其与肾脏水通道蛋白2 基因表达的相关性。方法　应用左冠状动脉结扎制备慢性心力衰竭大鼠模型 ,术后第 9周应用Western蛋白印迹测定尿液水通道蛋白2 浓度和肾脏水通道蛋白2 表达水平。结果　慢性心力衰竭大鼠和对照组大鼠尿液水通道蛋白2 吸光度比分别为 (36 5 5± 10 2 9) %和 (98 5± 47 6 ) %(P <0 0 1) ,心力衰竭大鼠尿液水通道蛋白2 浓度与肾脏水通道蛋白2 基因表达水平有较好的相关性(r=0 89,P <0 0 1)。结论　慢性心力衰竭大鼠模型尿液水通道蛋白2 浓度明显增加     

MAL decreases the internalization of the aquaporin-2 water channel

Body water homeostasis depends critically on the hormonally regulated trafficking of aquaporin-2 (AQP2) water channels in renal collecting duct epithelial cells. Several types of posttranslational modifications are clearly involved in controlling the distribution of AQP2 between intracellular vesicles and the apical plasma membrane. Little is known, however, about the protein interactions that govern the trafficking of AQP2 between these organelles. MAL is a detergent-resistant membrane-associated protein implicated in apical sorting events. We wondered, therefore, whether MAL plays a role in the regulated trafficking of AQP2 between intracellular vesicles and the apical surface. We find that AQP2 and MAL are coexpressed in epithelial cells of the kidney collecting duct. These two proteins interact, both in the native kidney and when expressed by transfection in cultured cells. The S256-phosphorylated form of AQP2 appears to interact more extensively with MAL than does the water channel protein not phosphorylated at this serine. We find that MAL is not involved in detergent-resistant membrane association or apical delivery of AQP2 in LLC-PK(1) renal epithelial cells. Instead, MAL increases the S256 phosphorylation and apical surface expression of AQP2. Furthermore, internalization experiments show that MAL induces surface expression of AQP2 by attenuating its internalization. Thus, the involvement of MAL in the cell surface retention of apical membrane proteins could play an important role in regulated absorption and secretion in transporting epithelia.     

Short-chain ubiquitination mediates the regulated endocytosis of the aquaporin-2 water channel

To regulate mammalian water homeostasis, arginine-vasopressin (AVP) induces phosphorylation and thereby redistribution of renal aquaporin-2 (AQP2) water channels from vesicles to the apical membrane. Vice versa, AVP (or forskolin) removal and hormones activating PKC cause AQP2 internalization, but the mechanism is unknown. Here, we show that a fraction of AQP2 is modified with two to three ubiquitin moieties in vitro and in vivo. Mutagenesis revealed that AQP2 is ubiquitinated with one K63-linked chain at K270 only. In Madin-Darby canine kidney cells, AQP2 ubiquitination occurs preferentially when present in the apical membrane, is transiently increased with forskolin removal or PKC activation, and precedes its internalization. Internalization kinetics assays with wild type (wt) and ubiquitination-deficient (K270R) AQP2 revealed that ubiquitination enhances AQP2 endocytosis. Electron microscopy showed that a translational fusion of AQP2 with ubiquitin (AQP2-Ub) localized particularly to internal vesicles of multivesicular bodies (MVBs), whereas AQP2-K270R largely localized to the apical membrane, early endosomes, and the limiting membrane of MVBs. Consistent with this distribution pattern, lysosomal degradation was extensive for AQP2-Ub, low for AQP2-K270R, and intermediate for wt-AQP2. Our data show that short-chain ubiquitination is involved in the regulated endocytosis, MVB sorting, and degradation of AQP2 and may be the mechanism used by AVP removal and PKC-activating hormones to reduce renal water reabsorption. Moreover, because several other channels are also (short-chain) ubiquitinated, our data suggest that ubiquitination may be a general mediator for the regulated endocytosis and degradation of channels in higher eukaryotes.     

Urinary excretion of free cortisol in impaired renal function.

Total cortisol and free, non protein-bound coritsol in plasma and urinary excretion of unconjugated free cortisol were measured during iv infusion of cortisol at varying dose rates in eight patients with impaired renal function. The results showed that free urinary cortisol decreased with decreased glomerular filtration rate (GFR), also compared to free cortisol level in plasma. An increase in free cortisol in plasma had no influence on GFR. It is concluded that determination of free urinary cortisol, otherwise useful in diagnosing Cushing's syndrome, may be of less value in patients with impaired renal function.     

The role of vasopressin in the impaired water excretion of myxedema

The plasma vasopressin response to acute water ingestion was evaluated in 20 patients with myxedema prior to definitive treatment and in eight of these same patients following therapy of their hypothyroidism. Vasopressin levels were elevated and failed to completely suppress following water ingestion in 15 subjects (75 per cent). Two hypothyroid patients with elevated plasma vasopressin levels (10 per cent) had a normal renal response to the water challenge suggesting partial end organ hormonal unresponsiveness. In three (15 per cent) of the five patients with suppressible vasopressin, water excretion was impaired indicating a nonvasopressin-mediated renal defect. In eight patients restudied after achievement of a euthyroid state, vasopressin inhibltion and urinary excretion were normal following the oral water load. Although intrinsic renal changes in the hypothyroid state may contribute to the observed defect in water diuresis, the present study suggests a role of endogenous vasopressin in this disorder.     

Urinary protein excretion in Type 2 diabetes with complications

This study examined the association between urinary markers of early diabetic nephropathy and non-renal diabetic complications in 946 patients with type 2 diabetes mellitus. The association with hypertension was also studied. Data on macrovascular complications (ischaemic heart disease, stroke, peripheral vascular disease) and microvascular complications (retinopathy, peripheral neuropathy) were obtained from case records and clinical examination. Urine samples collected were analysed for albumin, beta(2)-microglobulin, retinol-binding protein (RBP), and N-acetyl-beta-D-glucosaminidase (NAG). Results showed that urinary albumin, RBP and beta(2)-microglobulin levels were higher in patients with macro- and/or microvascular complications, compared to those without. NAG levels were higher only in patients with both types of complications. A higher proportion of patients with complications had abnormally raised urinary protein and enzyme levels, compared to those without. Patients with associated hypertension had higher urinary levels of albumin and beta(2)-microglobulin, regardless of whether complications were present or not. RBP excretion was, however, markedly higher only in patients with microvascular complications, whereas hypertension did not influence NAG excretion. Urine albumin and RBP excretion were predictive of microvascular, as well as both macrovascular and microvascular complications, whereas NAG excretion was predictive of macro- and microvascular complications. These findings could mean that increased urinary protein and enzyme excretion were associated with more severe disease in these patients.     

Urinary albumin excretion is associated with impaired flow- and nitroglycerin-mediated brachial artery dilatation in hypertensive adults

We investigated whether the urinary albumin/creatinine ratio (UACR), a measure of albuminuria, is associated with non-invasive measures of arterial function in hypertensive adults without known coronary heart disease (CHD) or stroke. UACR was measured in the first voided morning urine sample in 469 non-Hispanic white hypertensive individuals (mean age 62.2+/-9.8 years, 41% men) belonging to hypertensive sibships. High-resolution ultrasonography of the brachial artery was used to assess flow-mediated dilatation (FMD)--an endothelium-dependent response--and nitroglycerin-mediated dilatation (NMD)--an endothelium-independent response. Because of skewed distribution, UACR was log transformed after addition of 0.1. The association of log (UACR+0.1) with FMD and NMD, before and after adjustment for CHD risk factors, serum creatinine, and hypertension medication and statin use was assessed using linear regression analyses. In univariable analyses, variables associated with lower FMD were greater age, male sex, history of smoking, lower high-density lipoprotein (HDL) cholesterol, higher serum creatinine and higher log (UACR+0.1); variables associated with lower NMD were greater age, male sex, higher systolic blood pressure, lower HDL cholesterol, higher serum creatinine and higher log (UACR+0.1). In separate stepwise multivariable regression analyses that adjusted for conventional CHD risk factors, serum creatinine and hypertension medication and statin use, higher log (UACR+0.1) was associated with lower brachial artery FMD (P=0.035) and NMD (P=0.0002). These findings highlight the association of increased urinary albumin excretion with impaired vascular reactivity in hypertensive individuals.