Augmentation of intrarenal angiotensin II levels in uninephrectomized aldosterone/salt-treated hypertensive rats; renoprotective effects of an ultrahigh dose of olmesartan.

Recent studies have suggested that aldosterone plays a role in the pathogenesis of renal injury. In this study, we investigated whether local angiotensin II (Ang II) activity contributes to the progression of renal injury in aldosterone/salt-induced hypertensive rats. Uninephrectomized rats were treated with 1% NaCl in a drinking solution and one of the following combinations for 6 weeks: vehicle (2% ethanol, s.c.; n=9), aldosterone (0.75 mug/h, s.c.; n=8), aldosterone+Ang II type 1 receptor blocker olmesartan (10 mg/kg/day, p.o.; n=8), or aldosterone+olmesartan (100 mg/kg/day, p.o.; n=9). Aldosterone/salt-treated hypertensive rats exhibited severe proteinuria and renal injury characterized by glomerular sclerosis and tubulointerstitial fibrosis. Aldosterone/salt-induced renal injury was associated with augmented expression of angiotensin converting enzyme and Ang II levels in the renal cortex and medullary tissues. Renal cortical and medullary mRNA expression of transforming growth factor-beta (TGF-beta) and connective tissue growth factor (CTGF) as well as the collagen contents were increased in aldosterone/salt-treated hypertensive rats. Treatment with olmesartan (10 or 100 mg/kg/day) had no effect on blood pressure but attenuated proteinuria in a dose-dependent manner. Olmesartan at 10 mg/kg/day tended to decrease renal cortical and medullary Ang II levels, TGF-beta and CTGF expression, and collagen contents; however, these changes were not significant. On the other hand, an ultrahigh dose of olmesartan (100 mg/kg/day) significantly decreased these values and ameliorated renal injury. These data suggest that augmented local Ang II activity contributes, at least partially, to the progression of aldosterone/salt-dependent renal injury.     

Neural control of urinary sodium excretion during hypertonic NaCl load in conscious rabbits: role of renal and hepatic nerves and baroreceptors

We examined responses of renal nerve activity, urine flow, and urinary Na+ excretion to a hypertonic NaCl infusion in chronically instrumented conscious rabbits with unilateral renal denervation. The intravenous infusion of 20% NaCl, at 0.2 ml/min for 30 min, increased plasma osmolality by 27 +/- 5 mOsm/kg, and plasma Na+ by 16 +/- 3 mEq/l, and decreased hematocrit by 5 +/- 1%. These changes were accompanied by a marked decrease in renal nerve activity by 82 + 7%. Urine flow and urinary Na+ excretion increased gradually and peaked at the end of infusion. The innervated kidney excreted 23.3 +/- 3.3 ml urine and 5.5 +/- 0.7 mEq Na+ for the subsequent 60 min. However, the contralateral denervated kidney excreted only 9.5 +/- 2.0 ml urine and 2.2 +/- 0.6 mEq Na+; these values were significantly less than those of the innervated kidney. To examine the role of the sinoaortic and cardiopulmonary baroreceptors and the hepatic nerves in the response of renal nerve activity to the hypertonic NaCl infusion, renal nerve activity was examined in conscious rabbits with sinoaortic baroreceptor denervation (SAD) plus vagotomy and/or section of the anterior and posterior hepatic nerves (SAPH). In rabbits with SAD plus vagotomy or SAPH, the NaCl infusion also decreased renal nerve activity. After combining SAPH and SAD plus vagotomy, the decrease in renal nerve activity was completely blocked. These results indicate that hypertonic NaCl infusion elicits a marked decrease in renal nerve activity which is mediated predominantly by sinoaortic and cardiopulmonary baroreflexes and the hepatic nerves, and that the decrease in renal nerve activity plays an important role in the augmentation of renal function.     

Reflex control of renal nerve activity originating from the osmoreceptors in the hepato-portal region.

The reflex effects of hepatic osmoreceptors on the renal sympathetic nerve activity (RNA) were studied in 30 pentobarbital anesthetized, vagotomized and sino-aortic baroreceptor denervated (SAD + VD) rabbits. The changes in mean arterial pressure (MAP), heart rate (HR), and RNA were examined when 9% NaCl, 6.5% LiCl or 50% glucose solution was infused into the hepatic portal vein at a rate of 0.15 ml/kg/min for 10 min. Infusion of 9% NaCl solution into the hepatic portal vein increased the plasma osmolality by 10.8 +/- 1.0 mOsmol/kg from the control level in the blood of the hepatic portal vein and by 2.8 +/- 2.0 mOsmol/kg from the control level in the systemic blood. MAP was significantly elevated by 10.2 +/- 5.0 mmHg but HR did not change with hepatic portal infusion of 9% NaCl solution. Intraportal infusion of 9% NaCl solution significantly decreased the RNA by 28.6-34.2% from the control level, 6.5% LiCl solution by 28.6 +/- 4.7%, and 50% glucose solution by 26.2 +/- 3.0%. Femoral arterial infusion of hypertonic NaCl solution, however, did not evoke any significant change in RNA in SAD + VD rabbits. These findings suggest that increases in osmolality and NaCl concentration in the systemic circulation do not result in a decrease of RNA. Furthermore, after section of the anterior and posterior plexus of the hepatic nerve, hepatic portal infusion of hypertonic NaCl solution elicited no change in RNA. The present data indicate that an increase in osmolality in the hepatic portal venous blood results in a reflex decrease of RNA. This reflex may be important for restoration of a postprandial increase in osmolality.     

ENDOSCOPIC SUBMUCOSAL DISSECTION FOR RECURRENT GASTRIC TUMORS

Endoscopic resection has been accepted as the standard treatment for intramucosal gastric tumors of differentiated type. However, the indication was limited to small tumors to achieve en bloc resection and prevent local recurrence in cases of conventional endoscopic mucosal resection (EMR) such as the strip biopsy and the cap technique. To avoid multi‐fragmental resection, we have developed endoscopic submucosal dissection (ESD) as a new endoscopic resection technique. ESD is a remarkable technique, because we make it possible to remove the lesions en bloc regardless of size, shape, coexisting ulcer, and location. However, it is difficult or impossible to resect recurrent tumors en bloc in conventional EMR owing to hard fibrosis, and some patients need laparotomy. Using ESD, we can dissect the submucosal layer as we directly look at the submucosa, and remove the lesion safely and reliably even in cases of hard fibrosis. The key to treatment of recurrent tumors in ESD are as follows: (i) using enough submucosal injection solution (we use a mixture of Glyceol and 1% 1900 kDa hyaluronic acid preparation); (ii) incising the mucosa without fibrosis; (iii) understanding characteristics of various cutting devices, and changing other devices in difficult situations. In these ways we can remove the majority of the recurrent tumors en bloc. Hence, we consider that ESD is a very effective treatment which achieves excellent en bloc and complete resection rates and enables patients with intramucosal gastric tumors to a recurrent‐free survival even in recurrent tumors.     

Preventive effects of intermittent administration of human parathyroid hormone on steroid-induced osteopenia in rats

The purpose of this study was to determine the preventive effect of intermittent administration of human parathyroid hormone (h-PTH) on bone change in steroid-treated rats; this was done by histomorphometric and biochemical analysis. Seven-month-old female Wistar rats were divided into four groups; in-each of the four groups one subgroup was treated for 4 weeks and one for 8 weeks. The groups consisted of: untreated controls, a steroid group (receiving prednisolone), a steroid + PTH group (predniso-lone and h-PTH administered simultaneously), and a steroid + PTH vehicle group. Prednisolone (2.5 mg/kg) and h-PTH (1–34) (6.0 μg/kg) were administered six times a week during the experimental period. At necropsy, bilateral tibiae were collected: one was used for preparing undecalcified sections after Villanueva bone staining, and the other for decalcified tartrate-resistant acid phosphatase (TRAP) stained sections. Biochemical analysis showed that steroids increased urinary calcium at the 8th week; however, such bone metabolic markers as serum 1,25-(OH)₂D and urinary deoxypyridinoline did not change in any treatment group. Histomorphometrically, steroid-induced osteopenia was established at the 8th week by inhibition of both bone formation and bone resorption. The simultaneous intermittent administration of PTH plus steroid, however, increased both bone formation and bone resorption, resulting in increases in bone volume beginning at 4 weeks. These results suggest that the simultaneous intermittent administration of PTH with steroid prevents steroid-induced low-turnover osteopenia by stimulating bone turnover.     

Distribution characteristics of immunosuppressants FK506 and cyclosporin A in the blood compartment

Using two representative immunosuppressants, FK506 (FK) and cyclosporin A (CyA), of which the mechanism of pharmacological action is the same although there is a great difference in the pharmacological intensity, the distribution characteristics were studied in both in vivo and in vitro experiments using rat, dog, and human blood. Blood samples were fractionated by means of sedimentation in Ficoll-Paque®, and the drug contents in the diluted plasma fraction, erythrocyte fraction, and lymphocyte fraction were measured by an HPLC method. FK distributes to the lymphocyte fraction to a level about three times greater than that of CyA, while CyA distributes to the erythrocyte fraction to a level ten times that of FK. The distribution pattern of these fractions was independent of the drug concentration and species after correcting the drug concentration in each fraction with the blood drug concentration. The uptakes of FK and CyA in the isolated lymphocytes obtained from the rat spleen and human peripheral blood were also studied. The amount of FK taken up by the spleen lymphocytes is five times greater than that of CyA. In the case of the uptake study using human peripheral blood lymphocytes, the concentration of FK in the lymphocyte is 100-fold higher than that of CyA. This difference in the lymphocyte level between the two immunosuppressants is thought to be one of the reasons why FK is more potent than CyA, a difference of about 100-fold in the in vitro pharmacological study and about tenfold in the in vivo organ transplantation experiments.     

A case of retroperitoneal Hodgkin's disease with dysuria

A case of retroperitoneal Hodgkin's disease with dysuria is reported. A 56-year-old man visited our hospital with the complaints of dysuria and lower abdominal mass. On physical examination, an unmovable hard smooth mass of fist size was palpable in the lower abdomen and prostate was slightly swelling by rectal digital examination. Excretory urography demonstrated medial deviation of left lower ureter and bladder deformity. Retrograde urethrocystography showed deviation and compression of prostatic urethra. On CT, tumors were composed of several round masses, which surrounded the left common iliac artery on angiography. Surgical extirpation was carried out and histological examination revealed Hodgkin's disease. As postoperative treatment, chemotherapy with cyclophosphamide, adriamycin, vincristine and prednisolone was performed, and 30 months after the operation the patient was asymptomatic.     

Improved affinity of a human anti-Entamoeba histolytica Gal/GalNAc lectin Fab fragment by a single amino acid modification of the light chain

We previously produced, in Escherichia coli, a human monoclonal antibody Fab fragment, CP33, specific for the galactose- and N-acetyl-D-galactosamine-inhibitable lectin of Entamoeba histolytica. To prepare antibodies with a higher affinity to the lectin, recombination PCR was used to exchange Ser91 and Arg96 in the third complementarity-determining region of the light chain with other amino acids. The screening of 200 clones of each exchange by an indirect fluorescent antibody test showed that 14 clones for Ser91 and nine clones for Arg96 reacted strongly with E. histolytica trophozoites. Sequence analyses revealed that the substituted amino acids at Ser91 were Ala in five clones, Gly in three clones, Pro in two clones, and Val in two clones, while the amino acid at position 96 was substituted with Leu in three clones. The remaining eight clones exhibited no amino acid change at position 91 or 96. These mutant Fab fragments were purified and subjected to a surface plasmon resonance assay to measure the affinity of these proteins to the cysteine-rich domain of lectin. Pro or Gly substitution for Ser91 caused an increased affinity of the Fab, but substitution with Ala or Val did not. The replacement of Arg96 with Leu did not affect affinity. These results demonstrate that modification of antibody genes by recombination PCR is a useful method for affinity maturation and that amino acid substitution at position 91 yields Fabs with increased affinity for the lectin.