Immunohistochemical localization of glomerular basement membrane antigens in various renal diseases

Summary The immunofluorescent localization of glomerular basement membrane (GBM) antigens was examined in 52 specimens from normal kidneys and in various renal diseases using antisera to human GBM HGBM), IV type collagen (IV Col) and P3 antigen, a rat nephritogen. Anti-HGBM serum normally stained the GBM and the mesangium in a restrictive pattern, anti-IV Col serum stained the GBM and the mesangium in a wider pattern and anti-P3 serum stained only the GBM. In mesangial proliferative glomerulonephritis, including IgA nephropathy pathy and Henoch-Schönlein nephritis, the widened mesangial areas were stained with anti-HGBM and anti-IV Col sera. In membranous nephropathy, the punched-out lesions of thickened GBM were demonstrated with the three antisera in moderate cases and a double linear distribution with fine granulation with anti-HGBM and anti-IV Col sera were revealed in one severe case. In membranoproliferative glomerulonephritis, the expanded mesangium and thickened capillary walls were stained with anti-HGBM and anti-IV Col sera, while the outer line of glomerular capillary walls was only positive with anti-P3 serum. In crescentic glomerulonephritis, the collapsed glomerular tufts were stained normally with anti-HGBM and anti-P3 sera and weakly with anti-IV Col serum. In diabetic nephropathy, anti-HGBM serum stained the GBM in a double linear distribution without reacting with the expanded mesangium; anti-IV Col serum stained the mesangium and the GBM in a less clear double linear fashion while anti-P3 serum stained the GBM as single line. Thin membrane disease and Alport's syndrome had normal reactivity with all antisera. However, in one case of Alport's syndrome anti-HGBM and anti-P3 sera stained the GBM in a focal and segmental pattern, while normal staining with anti-IV Col serum was found. In lesions with adhesions and crescents the staining was positive for HGBM and IV Col and negative for P3; obsolescent glomeruli were stained with anti-HGBM and anti-P3 sera, and had diminished staining with anti-IV Col serum.The identification of the various structural glomerular antigens is useful in the classification of certain types of glomerular diseases. Further insight into the mechanisms underlying these conditions may be obtained in this way.     

Clinical significance of determination of serum RNase activities in patients with eosinophilia--II. Measurements using polyuridylic acid and polycytidylic acid as substrates

The activities of RNase (RNase-U and RNase-C) were determined in the serum and leukocytes of 277 patients with 14 cases of various kinds of eosinophilia (not less than 10(3)/microliters), 28 cases of chronic myelocytic leukemia (CML), using polyuridylic acid and polycytidylic acid as synthetic substrates according to the method of Raddi et al. Serum RNase-U activity, serum RNase-C activity and the activity ratio (U/C x 10(-3)) were 55 +/- 14 U, 1,280 +/- 235 U and 44 +/- 11 (mean +/- SD), 196 +/- 137, 1,992 +/- 1,134 U and 97 +/- 38, and 110 +/- 50 U, 1,854 +/- 625 U and 65 +/- 13 for normal subjects, eosinophilia and CML (untreated), respectively. U/C ratio in eosinophilia and CML (untreated) showed a highly significant positive correlation (p less than 0.001) with peripheral eosinophil count; the activity of serum RNase-U per cells in the supernatant of eosinophil homogenate rose significantly (p less than 0.001) compared with that of lymphocytes or granulocytes. Besides, serum and eosinophil RNase-U had a similar optimal pH. These results suggested that serum RNase-U in eosinophilia originated mostly from eosinophils and its rise was correlated strongly with the increase in eosinophils.     

Suppressive effects of E3330, a novel quinone derivative,on tumor necrosis factor-α generation from monocytes and macrophages

E3330 {(2E)-3-[5-(2,3-dimethoxy-6-methyl-1,4-benzoquinoyl)]-2-nonly-2-propenoic acid}, a novel synthesized hepatoprotective compound, has suppressive effects on tumor necrosis factor- (TNF-) generation from monocytes/macrophagesin vitro. E3330 (1–100 M) reduced lipopolysaccharide (LPS, 10 mg/ml or 1g/ml)-induced TNF- generation from rat resident andPropionibacterium acnes (P. acnes)-elicited peritoneal macrophages, rat and human monocytes, rat Kupffer cells, and splenic mononuclear cells in a concentration-dependent manner. E3330 also (1–100 M) suppressed TNF- generation stimulated with egg-albumin immune complex in ratP. acnes-elicited peritoneal macrophages. Northern blot analysis showed that LPS-induced expression of TNF- messenger RNA (mRNA) in human blood monocytes was suppressed by E3330. These findings indicate that E3330 has a suppressive effect on TNF- generation from monocytes/macrophages, regardless of origin or species, and this effect is based in part on the suppression of TNF- mRNA expression.     

Topical application of FK506 (tacrolimus) ointment inhibits mite antigen-induced dermatitis by local action in NC/Nga mice

BACKGROUND: FK506 ointment (tacrolimus ointment, protopic) is a new drug therapeutically effective for patients with atopic dermatitis (AD). However, the mechanism of action of FK506 ointment on AD is not fully understood. METHODS: We examined the effect of FK506 ointment on mite antigen-induced dermatitis in NC/Nga mice. Clinical symptoms and ear thickness were recorded, and histopathological studies and in vitro analyses were performed. RESULTS: Topical application of FK506 ointment (0.03-0.3%) suppressed the development of dermatitis. In the lesional skin, both interleukin (IL)-4 and interferon (IFN)-gamma were detected, even though the IL-4+/IFN-gamma- T helper 2 (Th2) population was predominant in the regional lymph nodes (LNs). Topical application of FK506 treatment reduced the elevated level of both IL-4 and IFN-gamma in the skin, but did not decrease the expansion of the Th2 population in the LNs. CONCLUSIONS: Topical application of FK506 ointment suppresses dermatitis by inhibiting the activation of inflammatory cells locally, without systemic immune suppression, in this AD model.     

Effects of testosterone and prolactin on rat prostatic weight, 5alpha-reductase, and arginase.

Prostatic weights, 5alpha-reductase, and arginase activities were utilized as indexes for the effects of prolactin in short-term experiments in intact, hypophysectomized or castrated rats. Experiments were performed in which a dose-related response in the above parameters was obtained with testosterone administration in castrated mature and immature rats in order to evaluate the effects of simultaneously administered prolactin. This approach was necessitated by the failure of prolactin alone to affect the parameters listed in intact, castrated or hypophysectomized rats. It was shown that ovine prolactin may have an enhancing effect on the prostatic weight, 5alpha-reductase, and arginase activities, but that this effect is neither consistent nor striking when compared to that of testosterone.. Nevertheless, it is still possible that the long-term effects of prolactin, even if they are only of an enhancing quality, may play an important role in normal prostatic physiology and in abnormal states.     

Possible role of autoantibodies against nephrin in an experimental model of chronic graft-versus-host disease

Nephrin, a product of the NPHS1 gene, is a component of the slit diaphragms that are found between glomerular foot processes and is a crucial element for glomerular filtration barrier. Recently, nephrin has been focused in a number of studies of proteinuria development including various types of acquired glomerular diseases including minimal change nephrotic syndrome and membranous nephropathy. However, the precise role of nephrin in such acquired glomerular diseases is still unknown. To analyse the role of nephrin further, two kinds of anti-nephrin antibodies were raised in the rabbits and applied to an experimental mouse model of chronic graft-versus-host disease, in which (C57BL/10 x DBA/2) F1 mice developed clinically apparent severe proteinuria with significant glomerular lesions 7 weeks after parental DBA/2 cell transfer. Antibody-sandwich ELISA detected anti-nephrin antibodies during week 2 to week 6, with the peak at week 2 or week 4. Colocalization of nephrin and IgG on week 4, week 6, and week 8 was revealed by confocal microscopic analysis, suggesting that in situ immune complex formation with nephrin in glomerular lesion. Taken together, it seems to be suggested nephrin and its autoantibody have a certain role in the development of glomerular lesion in our model mice.     

In vivo release of cisplatin from a needle-type copolymer formulation implanted in rat kidney

Cisplatin, cis-dichlorodiamine platinum (II), was incorporated in a needle-type copolymer formulation (0.8 mm diameter, 6 mm long) by radiation-induced polymerization. The copolymer used was copoly(diethylene glycol dimethacrylate/polyethylene glycol #600 dimethacrylate, 80/20 vol%). This copolymer, containing 6 mg of cisplatin, was implanted into the kidney of adult male Wistar rats (420 +/- 20 g). A total of 70 d was required for 100% release of cisplatin in vivo. The kidney tissue surrounding the formulation was strongly necrotized by the action of cisplatin. Two layers of necrosis could be distinguished: necrotic tissue surrounding the formulation and necrobiotic tissue surrounding the necrotic tissue. The amount of necrotic tissue changed markedly over time, but no change was apparent in the amount of necrobiotic tissue. The maximal amounts of necrotized tissue were observed 14 d after implantation: 3100 microns and 600 microns thick for the necrotic and necrobiotic tissues, respectively.     

Simultaneous localization of histamine and factor VIII-related antigen in the endothelium of the human umbilical vein

Simultaneous immunoelectron microscopic localization of histamine and factor VII-related antigen was examined on the same ultrathin section of the endothelium of the human umbilical vein from full-term deliveries by means of the double-immunolabeling technique. Small gold particles demonstrating antibody reaction with histamine are preferentially located in the cytoplasmic matrix and organelles, especially in mitochondria and on the luminal membrane surface of the endothelial cells. The gold particles representing histamine immunoreactivity also located on some of Weibel Palade (WP) bodies. In contrast, large gold particles demonstrating factor VII-related antigen are concentrated preferentially on most WP bodies. Single labeling of either histamine or factor VIII-related antigen shows similar results to those of the double labeling. The present study indicates that some WP bodies are involved in storage of both factor VIII-related antigen and histamine, but others store factor VIII-related antigen only. This difference in contents of WP bodies may be induced during the development and maturation process of this inclusion. At any rate, it is reasonable to consider that WP bodies have important roles in both vascular tonus and hemostasis during the vascular obliteration.