Nasunin inhibits the lipopolysaccharide-induced pro-inflammatory mediator production in RAW264 mouse macrophages by suppressing ROS-mediated activation of PI3 K/Akt/NF-κB and p38 signaling pathways.

Nasunin is a major anthocyanin in eggplant peel. The purpose of this study was to examine the anti-inflammatory effects of nasunin in lipopolysaccharide (LPS)-stimulated RAW264 macrophages and to identify the molecular mechanisms underlying these effects. We found that nasunin reduced the LPS-induced secretion of tumor necrosis factor-α, interleukin-6 and nitric oxide, and expression of inducible nitric oxide synthase in a dose-dependent manner. Nasunin diminished LPS-induced nuclear factor-κB (NF-κB) activation by suppressing the degradation of inhibitor of κB-α and nuclear translocation of p65 subunit of NF-κB. Nasunin also attenuated the phosphorylation of Akt and p38, signaling molecules involved in pro-inflammatory mediator production. Moreover, nasunin inhibited the intracellular accumulation of ROS, leading to the suppression of NF-κB activation, Akt and p38 phosphorylation, and subsequent pro-inflammatory mediator production. These findings suggest that nasunin exerts an anti-inflammatory effect and this effect is mediated, at least in part, by its antioxidant activity.

Effect of a Ferric Citrate Formulation, a Phosphate Binder, on Oxidative Stress in Chronic Kidney Diseases-Mineral and Bone Disorder Patients Receiving Hemodialysis: A Pilot Study.

A ferric citrate formulation for treating hyperphosphatemia is a new therapeutic that not only suppresses the accumulation of phosphorus in patients with chronic kidney disease-mineral bone disorders (CKD-MBD), but also ameliorates anemia caused by iron deficiency. In contrast, it has been demonstrated that intravenous iron injection markedly increases oxidative stress. This study was designed to investigate the effect of a ferric citrate formulation on oxidative stress in CKD-MBD patients receiving hemodialysis therapy. Fifteen CKD-MBD patients undergoing dialysis were enrolled in this study. The patients were orally administered a ferric citrate formulation for 6 months. Their plasma phosphorus concentrations remained unchanged with the switch from other phosphorus adsorbents to the ferric citrate formulation. In addition, the ferric citrate formulation generally allowed for dose reduction of an erythropoiesis stimulating agent with an increased hematopoietic effect. The average values of plasma ferritin level increased after the introduction of a ferric citrate formulation, but did not exceed 100 (ng/mL). Interestingly, oxidative stress markers did not increase significantly, and anti-oxidative capacity was not significantly decreased at 6 months after the drug administration. Similarly, no change was observed in any inflammation markers. The ferric citrate formulation induces negligible oxidative stress in CKD-MBD patients receiving dialysis under the present clinical condition.

Expression of three isoforms of Na-K-2Cl cotransporter (NKCC2) in the kidney and regulation by dehydration.

Sodium reabsorption via Na-K-2Cl cotransporter 2 (NKCC2) in the thick ascending limbs has a major role for medullary osmotic gradient and subsequent water reabsorption in the collecting ducts. We investigated intrarenal localization of three isoforms of NKCC2 mRNA expressions and the effects of dehydration on them in rats. To further examine the mechanisms of dehydration, the effects of hyperosmolality on NKCC2 mRNA expression in microdissected renal tubules was studied. RT-PCR and RT-competitive PCR were employed. The expressions of NKCC2a and b mRNA were observed in the cortical thick ascending limbs (CAL) and the distal convoluted tubules (DCT) but not in the medullary thick ascending limbs (MAL), whereas NKCC2f mRNA expression was seen in MAL and CAL. Two-day dehydration did not affect these mRNA expressions. In contrast, hyperosmolality increased NKCC2 mRNA expression in MAL in vitro. Bradykinin dose-dependently decreased NKCC2 mRNA expression in MAL. However, dehydration did not change NKCC2 protein expression in membrane fraction from cortex and outer medulla and in microdissected MAL. These data show that NKCC2a/b and f types are mainly present in CAL and MAL, respectively. Although NKCC2 mRNA expression was stimulated by hyperosmolality in vitro, NKCC2 mRNA and protein expressions were not stimulated by dehydration in vivo. These data suggest the presence of the inhibitory factors for NKCC2 expression in dehydration. Considering the role of NKCC2 for the countercurrent multiplier system, NKCC2f expressed in MAL might be more important than NKCC2a/b.

Parathyroidectomy markedly reduces oxidative stress in a patient with primary hyperparathyroidism.

Parathyroidectomy for hyperparathyroidism has been associated with a survival benefit, but the mechanisms remain unclear. We are reporting on an 88-year-old female patient who had high serum calcium and intact parathyroid hormone levels associated with an enlarged parathyroid gland. A parathyroidectomy was performed due to a diagnosis of primary hyperparathyroidism. After the surgery, there was a marked decrease in the oxidative stress markers, such as the ratios of oxidized to unoxidized albumin and advanced oxidation protein products. These results suggest that parathyroidectomy reduces oxidative stress in patients with primary hyperparathyroidism, which may in part explain the reduced risk for cardiovascular and all-cause mortality after parathyroidectomy.

Effects of raloxifene on bone mineral metabolism in postmenopausal Japanese women on hemodialysis.

In addition to renal osteodystrophy, postmenopausal women on hemodialysis are at high risk for osteoporosis. Recent studies reported the effects of raloxifene, a selective estrogen receptor modulator for osteoporosis, in postmenopausal women. The present study evaluated the efficacy of raloxifene and its effects on bone mineral metabolism in postmenopausal Japanese patients on dialysis. In a prospective, multicentre study, 17 postmenopausal women on chronic hemodialysis with severe osteoporosis (bone mineral density [BMD]≤2 SD by bone densitometry) were treated with 60 mg/day raloxifene hydrochloride for 12 months. The study also included 10 age-matched control women. Vitamin D and calcium salts were not changed during the study. Intact parathyroid hormone (iPTH), serum calcium and phosphorus, and bone resorption marker (NTx) were measured, and BMD were determined by DEXA, at 0, 6, and 12 months after administration of raloxifene. The mean lumbar spine BMD at baseline was similar in the two groups. Raloxifene therapy (for 12 months) improved lumbar spine BMD (by 2.6%) in 53% of the patients, while 70% of the control group showed a reduction in BMD (by 4.0%). Raloxifene significantly decreased serum calcium and increased iPTH. Our results suggested that raloxifene improved trabecular BMD in postmenopausal Japanese women on hemodialysis. The effects of raloxifene on serum calcium and serum iPTH level suggest it improves bone resorption. Vitamin D and/or calcium salts should be added to raloxifene treatment to avoid secondary hyperparathyroidism.

Vitamin D receptor activator reduces oxidative stress in hemodialysis patients with secondary hyperparathyroidism.

Treatment with a vitamin D receptor activator (VDRA) has survival benefits probably related to its effects beyond the traditional role in mineral metabolism. We hypothesized that VDRA reduces oxidative stress in hemodialysis (HD) patients. To test this hypothesis, we investigated the effect of VDRA on the oxidative status of albumin in HD patients with secondary hyperparathyroidism. Eleven HD patients with secondary hyperparathyroidism were treated with calcitriol at an intravenous dose of 1.5 µg/week for four weeks. Serum intact parathyroid hormone, calcium and phosphorus were monitored and we measured the amount of oxidized albumin and albumin hydroperoxides form before and after calcitriol treatment. The ratio of oxidized to un-oxidized albumin was determined as a representative marker of oxidative stress. The radical scavenging activity of albumin was also evaluated. After four weeks of calcitriol therapy, there were no significant changes in serum intact parathyroid hormone, calcium, or phosphorus levels; however, the ratio of oxidized to un-oxidized albumin was markedly decreased and serum thiol content was significantly increased after calcitriol treatment. Furthermore, the radical scavenging activity of albumin was greater after calcitriol treatment compared with that of untreated albumin. Our data suggest that intravenous calcitriol treatment reduces oxidative stress and strengthens antioxidant defenses by inhibiting albumin oxidation in HD patients with secondary hyperparathyroidism.

Development of the ultra-miniaturized Inertial Measurement Unit WB3 for objective skill analysis and assessment in neurosurgery: preliminary results.

In recent years there has been an ever increasing amount of research and development of technologies and methodologies aimed at improving the safety of advanced surgery. In this context, several training methods and metrics have been proposed, in particular for laparoscopy, both to improve the surgeon's abilities and also to assess her/his skills. For neurosurgery, however, the extremely small movements and sizes involved have prevented until now the development of similar methodologies and systems. In this paper we present the development of the ultra-miniaturized Inertial Measurement Unit WB3 (at present the smallest, lightest, and best performing in the world) for practical application in neurosurgery as skill assessment tool. This paper presents the feasibility study for quantitative discrimination of movements of experienced surgeons and beginners in a simple pick and place scenario.

Comparison of sevelamer hydrochloride with colestimide, administered alone or in combination with calcium carbonate, in patients on hemodialysis.

Since hyperphosphatemia in hemodialysis patients can cause secondary hyperparathyroidism and promotes vascular calcification, serum phosphate (Pi) levels must be controlled by phosphate binders. Although sevelamer and colestimide are known as similar non-calcium, non-aluminum phosphate binders in hemodialysis patients, there are no studies that compare the effects of the two agents as either a monotherapy or in combination with calcium carbonate (CaCO3). We randomly allocated 62 hemodialysis patients with hyperphosphatemia to treatment with sevelamer (3.0 g/day) and colestimide (3.0 g/day). During the study, 35 subjects dropped out, leaving 13 in the sevelamer group and 14 in the colestimide group. After a 2-week CaCO3 washout, all subjects received the monotherapy for 4 weeks and then CaCO3 (3.0 g/day) was added for another 4 weeks. Serum corrected calcium levels tended to decrease in both groups during the washout period and monotherapy, but there was no significant difference between the two groups after the addition of CaCO3. Although the calcium x phosphorus product (Ca x P) in the two groups increased during the washout period, there was no significant change or difference between the two groups during monotherapy. However, the addition of CaCO3 significantly reduced serum Pi at Week 8 compared to that at Week 0 in both groups, and significantly lowered Ca x P only in the sevelamer group, but not in the colestimide group(.) In this short-term study, sevelamer and colestimide similarly ameliorated hyperphosphatemia, but the combination of sevelamer and CaCO3 was more effective than colestimide with CaCO3 in controlling the Ca x P product, and it may improve cardiovascular mortality in hemodialysis patients.

Association between long-term efficacy of cinacalcet and parathyroid gland volume in haemodialysis patients with secondary hyperparathyroidism.

Purpose. Secondary hyperparathyroidism with nodular hyperplasia is resistant to medical therapies. Cinacalcet is an effective treatment for severe secondary hyperparathyroidism. This multicentre retrospective study was designed to determine the long-term efficacy of cinacalcet in patients with nodular hyperplasia, the advanced type of parathyroid hyperplasia. Subjects and methods. The study subjects were 20 haemodialysis patients with secondary hyperparathyroidism. Patients with ultrasonographically confirmed large parathyroid glands (volume >0.5 cm(3)) were considered to have nodular hyperplasia (n = 8). Cinacalcet was started at the dose of 25 mg/day and titrated up to 100 mg/day to achieve the target intact-parathyroid hormone (iPTH) level of <250 pg/ml. Serum iPTH, corrected calcium, serum phosphorus, calcium × phosphorus product were measured and compared over the 48-week period of treatment with cinacalcet in all 20 patients and over 120 weeks in 6 of the patients (2 with nodular hyperplasia and 4 with non-nodular hyperplasia). We also examined the achievement rate of K/DOQI guideline treatment targets. The dosages of vitamin D preparation, sevelamer hydrochloride and calcium- containing phosphate binder were adjusted for the above target values. Results. iPTH levels were significantly lower at 48 weeks in both groups. However, corrected calcium levels, serum phosphorus levels and calcium phosphorus products were within the target values in the non-nodular hyperplasia group (n = 12), while the target value could not be achieved in the nodular hyperplasia group. In the long-term follow-up group, the levels of iPTH, corrected calcium, serum phosphorus and calcium × phosphorus products were significantly higher in nodular hyperplasia than in non-nodular hyperplasia. Conclusion. Our study suggests that cinacalcet lacks long-term efficacy in nodular hyperplasia, especially for controlling serum calcium and phosphorus levels.

The whole-PTH/intact-PTH ratio is a useful predictor of severity of secondary hyperparathyroidism.

Background. The newer parathyroid hormone (PTH) assay, whole-PTH, uses an antibody that binds the region harbouring the first amino acid, making it specific for the complete molecule, 1-84-PTH. Especially among dialysis patients, it has been reported that the level of whole-PTH can be calculated as ∼60% of their intact-PTH value. In addition, since 1-84-PTH is part of intact-PTH, the whole-PTH/intact-PTH ratio should not theoretically exceed 1. However, an abnormally high 1-84-PTH/intact-PTH ratio is reported in a few patients with parathyroid carcinoma, primary hyperparathyroidism and secondary hyperparathyroidism. In this study, we examined the correlation between the 1-84-PTH/intact-PTH ratio and the severity of hyperparathyroidism in patients on haemodialysis (HD). Patients and methods. The study population comprised 196 HD patients (males 113, females 83, age 67.4 ± 13.6 years, HD period 8.1 ± 7.3 years; mean ± SD). The whole-PTH/intact-PTH ratio was compared in patients with high PTH levels (intact-PTH ≥300 pg/ ml; high PTH group, n = 32), moderate PTH levels (intact-PTH >150-<300 pg/ml; moderate PTH group, n = 50) and low PTH levels (intact-PTH <150 pg/ml; low PTH group, n = 114). The ratio was also compared in 25 patients with at least one enlarged gland >0.5 cm(3) suggesting nodular hyperplasia, as determined by power Doppler ultrasonography (hyperplasia group) with seven patients without enlarged gland (non-hyperplasia group) and six patients who had undergone total parathyroidectomy (post-PTx group). Results. The whole-PTH/intact-PTH ratio of the high PTH group (0.68 ± 0.1) was significantly higher than those of the moderate (0.61 ± 0.1, P < 0.001) and low (0.52 ± 0.1, P < 0.001) groups. Moreover, the ratio was significantly higher in the hyperplasia group (0.70 ± 0.1) than those in the non-hyperplasia group (0.59 ± 0.1, P < 0.05) and post-PTx group (0.456 ± 0.12, P < 0.001). Conclusions. The whole-PTH/intact-PTH ratio correlated with the severity of hyperparathyroidism. Our results suggest that the ratio might be a useful predictor of severity of secondary hyperparathyroidism in HD patients.

Clinical usefulness of oral combination chemotherapy of 5'-deoxy-5-fluorouridine (5'-DFUR) and cyclophosphamide for metastatic breast cancer.

BACKGROUND: It has been reported that 5'-deoxy-5-fluorouridine (5'-DFUR), the pro-drug of 5-FU, is effective treatment for breast cancer that express thymidine phosphorylase (dThdPase). Since oral cyclophosphamide (CPA) induces dThdPase, a synergistic effect can be expected by combining CPA with 5'-DFUR. We evaluated the usefulness of combination chemotherapy using CPA and 5'-DFUR in patients with relapsed breast cancer in this prospective phase II study. METHODS: Patients with relapsed, advanced breast cancer with evaluable lesions were given 5'-DFUR at 800 mg/day/body and CPA at 100 mg/day/body for 2 weeks, then underwent 2 weeks of drug withdrawal. This was considered one course of treatment. It was repeated until progressive disease (PD) was confirmed. The lesions were evaluated according to UICC criteria and compared with regard to the clinical status. RESULTS: Sixty-four patients with relapsed, advanced breast cancer were registered. Complete response (CR) was seen in 7 patients, partial response (PR) in 12 patients, no change (NC) in 25 patients, of whom 11 achieved long NC with the effect lasting for more than 6 months, and PD was seen in 20 patients. The response rate was 29.7%. The total number of CR, PR, and long NC cases was 30, which comprise-46.9% of the total 64 cases (the clinical benefit rate). As for adverse events, hematological toxicities were seen in 9 patients, with grade 3 toxicits was seen in 1 patient. All other adverse events were grade 1 or 2. CONCLUSION: For those patients who achieved an effect more than NC, it was possible to continue the therapy for an average of 53 weeks. This treatment method is worth considering for patients who have metastatic breast cancer, that is not life threatening.

Combination therapy of intravenous maxacalcitol and percutaneous ethanol injection therapy lowers serum parathyroid hormone level and calcium x phosphorus product in secondary hyperparathyroidism.

BACKGROUND: Percutaneous ethanol injection therapy (PEIT) is an alternative treatment for secondary hyperparathyroidism (SHPT). Although maxacalcitol has been recently developed as a new vitamin D3 and its efficacy is anticipated in SHPT, there are only few reports on the usefulness of combination therapy of intravenous maxacalcitol and selective PEIT. METHODS: The study population comprised 10 hemodialysis patients (6 males and 4 females, mean age; 51.5 +/- 13.5 years, mean HD period 13.7 +/- 3.5 years), with high intact-PTH level (>400 pg/ml) and 1 or 2 enlarged parathyroid glands detected by power Doppler ultrasonography. Intravenous maxacalcitol therapy commenced one week after PEIT at 15 microg/week. The effect of combination therapy was monitored by measuring intact-PTH, serum Ca and P, bone metabolic markers, parathyroid gland volume and bone mineral density, prior to and at 6 and 12 months after PEIT. RESULTS: Successful control of intact-PTH, bone metabolic markers and parathyroid gland volume was achieved using the combination therapy. Serum P and CaxP product were significantly decreased 12 months after PEIT. The mean values of serum intact-PTH, P and CaxP product fulfilled all of the K/DOQI guidelines at 12 months after PEIT. None of the patients developed complications related to PEIT-maxacalcitol therapy during 12 months following PEIT. CONCLUSION: Combination therapy of intravenous maxacalcitol therapy and selective PEIT is safe and effective for the treatment of refractory SHPT. This combination therapy results in suppression of PTH secretion, regression of parathyroid hyperplasia and the control of CaxP product, which may prevent calcific uremic arteriolopathy in dialysis patients.

Application of neural network to humanoid robots-development of co-associative memory model.

We have been studying a system of many harmonic oscillators (neurons) interacting via a chaotic force since 2002. Each harmonic oscillator is driven by chaotic force whose bifurcation parameter is modulated by the position of the harmonic oscillator. Moreover, a system of mutually coupled chaotic neural networks was investigated. Different patterns were stored in each network and the associative memory problem was discussed in these networks. Each network can retrieve the pattern stored in the other network. On the other hand, we have been developing new mechanisms and functions for a humanoid robot with the ability to express emotions and communicate with humans in a human-like manner. We introduced a mental model which consisted of the mental space, the mood, the equations of emotion, the robot personality, the need model, the consciousness model and the behavior model. This type of mental model was implemented in Emotion Expression Humanoid Robot WE-4RII (Waseda Eye No.4 Refined II). In this paper, an associative memory model using mutually coupled chaotic neural networks is proposed for retrieving optimum memory (recognition) in response to a stimulus. We implemented this model in Emotion Expression Humanoid Robot WE-4RII (Waseda Eye No.4 Refined II).

Efficacy of percutaneous ethanol injection therapy for secondary hyperparathyroidism in patients on hemodialysis as evaluated by parathyroid hormone levels according to K/DOQI guidelines.

Secondary hyperparathyroidism (SHPT) is a major complication of hemodialysis patients. Recently, percutaneous ethanol injection therapy (PEIT) has become a useful alternative treatment to parathyroidectomy (PTx). In this study, we evaluate the usefulness of PEIT for SHPT according to Kidney Disease Outcomes Quality Initiative (K/DOQI) guidelines. We studied 28 patients on hemodialysis with high intact-PTH (>400 pg/mL) and one to four swollen parathyroid glands detected by power Doppler ultrasonography. They were classified into Group 1 (N = 16), with 1 or 2 swollen glands, Group 2 (N = 5), with 3 or 4 swollen glands, and Group 3 (N = 7), high-risk patients for PTx. We compared serum intact-PTH levels 1 year after PEIT according to K/DOQI guidelines among these groups. We also evaluated the effectiveness of PEIT and PTx by comparing intact-PTH levels in 21 patients 1 year after PEIT (groups 1 and 2) with 11 patients after PTx. In Group 1, adequate intact-PTH levels were noted in 13 of 16 (81.2%) patients after PEIT, while 1 patient of 5 (20%) was achieved in Group 2, and 2 of 7 (28.6%) patients of Group 3. Adequate intact-PTH levels were attained in 14 of 21 (66.7%) patients of the PEIT group but only in 2 of 11 (18.2%) patients of the PTx group. Our results suggest that PEIT is a useful treatment for SHPT, especially in patients with one or two swollen glands. Through appropriate selection of patients for PEIT and correct injection of ethanol into the enlarged parathyroid gland, PEIT could accomplish better outcomes based on K/DOQI guidelines.

[Clinical courses of two male siblings on hemodialysis for Fabry disease ].

Fabry disease is an X-linked recessive disease resulting from a deficiency of the lysosomal hydrolase alpha-galactosidase A. In male patients with the classic hemizygous form, acroparesthesias, hypohidrosis, corneal opacities, and dysfunction of the heart, brain, and kidney are observed. Recently, it was reported that 0.5-1.2% of male chronic hemodialysis (HD) patients were diagnosed as having Fabry disease based on the measurement of alpha-galactosidase A activity. Fabry disease is thought to be an important cause of end-stage renal disease. There are a few reports of patients with Fabry disease on long-term HD. Here we report two male siblings with classical type Fabry disease on HD. They had acroparesthesias, and hypohidrosis. Their mother had severe heart failure due to a heterozygous form of Fabry disease. Case 1 is a 44-year-old male. He had mid-cerebral apoplexy at 30 years of age. He started maintenance HD in 2000. Remarkable left ventricular hypertophy and conduction disorders of the heart were found. In 2004, he collapsed and ventricular-tachycardia and severe hypoxic brain damage were found. Now his consciousness level has been in the range of 100 to 300 on the Japan Coma Scale. Case 2 is a 40-year-old male. He started maintenance HD in 1993. Malnutrition due to chronic diarrhea and severe ischemic change in the brain were found. In 1998, he had severe joint pain of shoulders and fingers with ectopic calcifications detected by X ray. The ectopic calcifications were extended to the whole body. In 2004, his dementia by ischemic change in the brain has rapidly progressed. In conclusion, cardiovascular complications, cerebrovascular manifestations, painful ectopic carcifications, and chronic diarrheas in our patients were considered to be specific symptoms of Fabry disease. Young HD patients with these symptoms will need to be examined for Fabry disease.

Ovarian non-small cell neuroendocrine carcinoma with paraneoplastic parathyroid hormone-related hypercalcemia.

Ovarian tumors associated with hypercalcemia due to ectopic secretion of parathyroid hormone (PTH) are extremely rare. A 33-year-old woman presented with a pelvic mass and profound hypercalcemia accompanied by an elevated serum level of PTH. Laparotomy demonstrated a left ovarian tumor that on histological examination was a neuroendocrine carcinoma of non-small cell type admixed with a component of endometrioid adenocarcinoma. After left salpingo-oophorectomy, the serum calcium and PTH levels normalized. The cells of the neuroendocrine carcinoma were positive for neuron-specific enolase, synaptophysin, chromogranin A, and PTH. Hypercalcemia and elevated serum PTH levels recurred during tumor relapse, and the patient died of disease 6 months postoperatively. This is the eleventh case of neuroendocrine carcinoma of non-small cell type associated with surface epithelial neoplasm of the ovary, and the first such tumor to be associated with hypercalcemia.

Infarction of mediastinal parathyroid gland causing spontaneous remission of secondary hyperparathyroidism.

Secondary hyperparathyroidism is a serious complication in long-term hemodialysis patients. The authors report on 2 patients on long-term hemodialysis who suffered from persistent secondary hyperparathyroidism due to missed mediastinal parathyroid gland after total parathyroidectomy with forearm autograft. Reoperation was planned. In both cases, severe hypocalcemia suddenly developed; serum parathyroid hormone (PTH) level decreased markedly after this episode. The serum calcium level increased gradually in response to administration of vitamin D and calcium carbonate, but serum PTH level remained low. A follow-up computed tomography scan showed that the formerly enlarged mediastinal parathyroid gland was markedly reduced in size. Moreover, a hot spot formerly detected by technetium 99m-MIBI (methoxy-isobutyl-isonitrile) scintigraphy in the mediastinum disappeared after this episode. The authors considered that necrosis of the enlarged ectopic parathyroid gland, probably due to infarction, resulted in hypocalcemia. To the authors' knowledge, this is the first case report of spontaneous mediastinal parathyroid autoinfarction after parathyroidectomy in hemodialysis patients.

Clinical usefulness of a new hepatitis C virus RNA extraction method using specific capture probe and magnetic particle in hemodialysis patients.

Hemodialysis patients are a high-risk group for hepatitis C virus (HCV) infection. Assessment of HCV infection using HCV-RNA assay among dialysis patients is important for the issue of safety and environmental protection. However, polymerase chain reaction (PCR)-based methods are unsuitable for analyzing samples from dialysis patients because the conventional centrifugal extraction method fails to eliminate heparin, a potent inhibitor of PCR. In this study, we evaluated the usefulness of a HCV-RNA extraction method using probes and magnetic particles for hemodialysis patients in comparison with the centrifugal method. The study population consisted of 17 HCV antibody-positive patients undergoing hemodialysis. These 17 patients consisted of 12 HCV carrier patients and five patients with past HCV infection. One hundred and two samples from these patients were measured using the centrifugal and magnetic methods. Moreover, we prepared five standards that included theoretically 5 KIU/mL of HCV. One was made from non-HD patient's serum and the other four were from hemodialysis patients' serum. These standards were measured using the two methods. False-negative results were not observed with the magnetic method, but were observed in five out of 102 samples with the centrifugal method. Studies using standard samples revealed that accurate HCV-RNA measurement is achieved using the magnetic method. In conclusion, the present study showed that this magnetic extraction method is a highly reproducible and reliable assay to obtain correct information about the presence of the infective virus itself in the hemodialysis setting. Precise identification of HCV-RNA using this specific method is considered to be useful in preventing HCV infection in hemodialysis units.

Gene regulation of renal-osmotic stress-induced Na-CL organic solute cotransporter.

BACKGROUND: Na- and Cl-dependent organic solute cotransporters participate in transporting neurotransmitters in the brain and organic osmolytes in the kidney. METHODS: We examined the intranephron localization and regulation of the renal osmotic-stress-induced Na-Cl organic solute cotransporter (ROSIT) mRNA expression using microdissected nephron segments from control and dehydrated rats and RT-PCR. To further know the mechanisms of gene regulation of ROSIT, microdissected proximal straight tubules (PST) were incubated in isotonic (290 mosm/kg H2O) or hyperosmotic (490- 1,090 mosm/kg H2O) solution. RESULTS: ROSIT mRNA was expressed predominantly in PST and to a lesser extent in cortical thick ascending limbs and cortical collecting ducts in control rats, and dehydration caused an increase in the expression in whole nephron segments. ROSIT mRNA in PST was decreased with time by incubation in isotonic solution. Incubation of PST in hypertonic solution by adding NaCl increased mRNA expression as early as 15 min (1.5- and 3-fold at 15 and 30 min, respectively). This stimulating effect of NaCl was largest at 890 mosm/kg H2O. Hypertonicity by mannitol or myoinositol also increased ROSIT mRNA expression. In contrast, hyperosmolality by urea reduced ROSIT mRNA expression. GAPDH mRNA expression, an internal standard, did not change by incubation in NaCl or mannitol solution. CONCLUSION: In summary, ROSIT mRNA expression was most abundant in PST in control and it was stimulated in whole nephron segments by dehydration. ROSIT mRNA expression in PST was stimulated by hypertonicity but not by urea. These data suggest that ROSIT may participate in the transport of amino acid under control conditions and organic osmolytes in dehydration.