[Chronic renal failure and pregnancy]. / Insuffisance rénale chronique et grossesse.

The women who are suffering from chronic renal failure in an advanced stage have a deficient fertility but they are not sterile. Hemodialysis has improved considerably the fertility of these patients. The aim of this study is to give the results of our experience, from 1990 to 1996, about pregnancies among the uremic patients, dialysed or no and to make a literature review about this subject. We have noticed that pregnancies in the dialysis patients are rare and their evolution is precarious. We have also noticed more miscarriage or pregnancy interruption. Complications are frequent. Mothers have a high risk of hemorrhagic accident (ablatio placentae), of anemia aggravation, of thrombosis of the vascular approach and a high risk of liver anomalies (gravidic cholestasis). The fetus suffers from the maternal anemia and from chronic hypoxia. He's threatened by hydramnios in the case of bad volemic supervision. The intra uterine delayed developement and the prematurity are usual. The absence of high blood pressure and a residual renal function are representing the favourable elements of the good march of pregnancy. A therapeutic intensification is necessary in order to lead this pregnancies to a viable term. The management is heavy not only for the nephrologic, the obstetrical and the neonatal physicians, but also for the patient who is the only one who can decide to continue or to interrupt the pregnancy. It seems better to inform the patient rather than to procure her abortion by proposing her an effective and inoffensive contraceptive method meanwhile to be pregnant after renal transplantation.

The effect of organ preservation solutions on kidney tubular and endothelial cells.

Organ preservation solutions have primarily been tested in whole organ animal models. In the current study, we have examined the effect of commonly used organ preservation solutions on both kidney tubular and endothelial cells. Primary human endothelial and kidney tubular cells were incubated at 4 degrees C in the following solutions: 0.9% saline (NS), EuroCollins (EC). University of Wisconsin (UW), or Hank's balanced salts with 5% polyethylene glycol (PEG). Cell viability was assessed by colorometric measurement of mitochondrial reduction of 3 (4,5-dimethylthiazol-2-yl)-2-,5-diphenyltetrazolium bromide (MTT) to purple 1-(4,5-dimethylthiazol-2-yl)-3,5-diphenylformazan. After hypothermic storage, cells were incubated at 37 degrees C in media with MTT, and the amount of reduced formazan present was quantified. Endothelial cells preserved in PEG displayed the best viability (P < 0.05). UW provided better cellular viability than EC or NS (P < 0.05). Control endothelial cells preserved in culture media at 37 degrees C displayed the highest absorbance values (P < 0.01). For kidney tubular cells, UW and PEG provided the best cellular protection (P < 0.05). Control kidney tubular cells cultured in complete media at 37 degrees C displayed the highest absorbance values (P < 0.01). Although the model presented here was not part of a truly morphological study, it may be more reliable for the rapid assessment of preservation-induced cell injury than models presented in previous morphological studies and may help in the development of improved preservation techniques.

Interleukin-1 and its naturally occurring antagonist in peritoneal dialysis patients.

We have assessed the levels of interleukin-1b (IL-1b) and interleukin-1 receptor antagonist (IL-1RA) in both serum and peritoneal dialysate effluents (PDE) from nineteen continuous ambulatory peritoneal dialysis patients (CAPD) without peritonitis and three CAPD patients with peritonitis. IL-1 beta and IL-1RA were tested using a specific ELISA immuno-assay. Fifteen normal healthy volunteers severed as control. The serum levels of IL-1RA in CAPD patients were significantly increased comparatively to their levels in healthy volunteers (p < 0.001). CAPD patients without peritonitis (stable patients) showed relatively low levels of IL-1RA in peritoneal dialysate effluents (114.4 +/- 85.1 pg/ml). Patients with peritonitis showed very high serum levels of IL-1RA at the onset of acute infection (4710 +/- 50 pg/ml). The levels of IL-1RA in PDE were very high during the onset of bacterial peritonitis (5744 +/- 254 pg/ml). The clinical recovery from peritonitis was characterized by a fall in IL-1RA in both serum and dialysate. Serum levels IL-1 beta showed a different pattern, it was not detectable in stable CAPD patients as well as in normal healthy volunteers. It was detectable only in serum of patients with peritonitis (10 +/- 0.8 pg/ml). Likewise, in most stable patients, IL-1 beta-PDE levels were not detectable, but substantial amounts can be detected in PDE during bacterial infection (80 +/- 15 pg/ml). The increase in serum and PDE levels of IL-1 beta during bacterial infection was very rapid, this cytokine disappeared in serum and PDE, 2 or 3 days before the clinical recovery.(ABSTRACT TRUNCATED AT 250 WORDS)

Human glomerular epithelial cells synthesize and secrete the third component of complement.

Complement proteins in serum are mainly synthesized by hepatocytes. Recently, many cell types have been reported to synthesize complement in various tissues. In this study, we report the synthesis and secretion of the third component of complement (C3) by cultured glomerular epithelial cells (GEC). Using reverse transcriptase polymerase reaction, we have found that GEC and whole kidney expressed the C3 mRNA for C3. By ELISA, we have found that C3 was secreted in culture supernatants harvested from cultured GEC. The secretion of C3 is regulated by proinflammatory cytokines (IL-1 beta, TNF-alpha and IL-6). IL-1 beta is shown to be the most potent stimulator of C3 secretion from GEC. The exact significance of C3 produced at glomerular site is not clear, but its upregulation by proinflammatory cytokines may suspect a role in local activation of complement which may lead to glomerular injury. We further studied the expression of C3 step regulatory proteins (membrane cofactor protein (MCP), decay-accelerating factor (DAF), CR-1 and CD59 (a terminal step regulatory protein) by cultured GEC. Treatment of GEC by proinflammatory cytokines IL-1 beta, TGF-beta, TNF-alpha and IL-6 did not modify the expression of MCP, DAF and CR-1 whereas an increase in the expression of CD59 could be observed after treatment with IL-1 beta and TGF-beta. These results indicate that the expression of these regulatory proteins is tissue specific and may be implicated in inflammatory processes.

Interleukin-6 and interleukin-6 receptor are expressed by cultured glomerular epithelial cells.

Interleukin-6 (IL-6) has been extensively studied in mesangial cells but little is known about the expression of this cytokine and its receptor in glomerular epithelial cells (GEC). IL-6 was detected in the culture supernatants of human GEC and its production was enhanced in time and dose dependent manner by lipopolysaccharide (LPS), interleukin-1 beta (IL-1 beta) and tumour necrosis alpha (TNF-alpha). Quiescent, serum-starved GEC did not express clearly IL-6 mRNA. Stimulation of cells with LPS, TNF-alpha or IL-1 beta resulted in an increase of detectable IL-6 mRNA. Interestingly, it was found that IL-6 induced its own mRNA attesting that this cytokine was secreted in autocrine fashion by GEC. GEC expressed IL-6 receptor (IL-6R) as demonstrated directly by the existence of IL-6R mRNA detected by northern blotting. Stimulation of GEC by pro-inflammatory mediators such as LPS increased the expression of IL-6R mRNA. The soluble form of IL-6 receptor (sIL-6R) was not detectable in the culture supernatants harvested from untreated or cytokine-treated cells. We investigated further, whether IL-6 may influence growth of cultured GEC. Incubation of GEC with recombinant (r) IL-6 resulted in a dose dependent increase in 3H thymidine incorporation indicating that IL-6 acts as an autocrine growth factor for GEC. We conclude that GEC are a potent source of IL-6, the local excessive expression of IL-6 and its receptor may play a substantive role in the regulation of processes which appear critical to the initiation of progressive glomerular disease such as cell proliferation.

Identification and characterization of membrane cofactor protein (CD46) in the human kidneys.

Membrane cofactor protein (MCP, CD46) is an integral protein that serves as a cofactor for factor I in inactivating C3b/C4b deposited on the same cell membrane as C3bi/C4c+C4d. This C3b/C4b inactivation is closely associated with self-protection of host cells from autologous complement attack. We have studied the distribution and properties of MCP in the normal human kidney by immunohistochemical and immunoblotting methods using monoclonal antibodies against MCP. MCP was predominantly expressed on the juxtaglomerular apparatus. Glomerular capillary walls, mesangial areas, and tubulus were also MCP positive. Glomerulus MCP was composed of two major bands of 45-65 kDa, which were similar to those of lymphocyte MCP. The proportion of the high and low molecular weight components in glomerulus MCP, however, was considerably different from that of lymphocyte MCP among the individual samples tested. Glomerular epithelial cells and mesangial cells from an individual having equal amounts of high and low molecular weight components in the lymphocytes were cultured separately and the properties of their MCP investigated. MCP in the mesangial cells and glomerular epithelial cells showed profiles in which the upper band was predominant. The results may explain the unique distribution of the high and low molecular weight forms in the glomerulus. These forms of MCP together with factor I were all capable of inactivating C3b to C3bi. Message analysis suggested that glomerular epithelial cells and mesangial cells synthesized a single species of mRNA of 4.2 kb from which the polymorphic MCP species were generated. Flow cytometric analysis suggested that MCP was minimal in mesangial cells. These results, taken together with the previous reports on the distribution of other complement regulatory proteins, infer that the distribution profile of MCP is rather similar to that of DAF but differs from those of CD59 and CR1 in the normal human kidney; this may reflect the differences between their roles or functional properties in renal tissue.

Interleukin-1-beta activation of cultured glomerular epithelial cells.

In crescentic glomerulonephritis, crescent formation involves the active participation of glomerular epithelial cells (GEC) and macrophages recruited to the glomerulus during the evolution of the disease. Cytokines derived from macrophages may affect many functions of GEC. In this study, we found that interleukin-1 beta (IL-1 beta) inhibited GEC growth (DNA synthesis and cell number) in vitro in a dose- and time-dependent manner. This effect was not mediated by tumor growth factor beta (TGF beta) which is a potent inhibitor of GEC growth in vitro. Treatment of GEC with various concentrations of IL-1 beta induced morphologic changes consisting in the loss of their cobblestone shape and acquisition of a fibroblast-like appearance. Moreover, IL-1 beta was shown to stimulate the expression of interleukin-6 (IL-6) by GEC. The increase in IL-6 secretion by GEC treated with IL-1 beta was observed at both the protein and mRNA levels. IL-1 beta also affected the metabolism of laminin in cultured GEC, inducing a dose-dependent increase in laminin production in culture supernatants harvested from GEC. Finally, we investigated the expression of MHC class II antigens and intercellular adhesion molecule-1 (ICAM-1) in GEC, and found that unstimulated GEC are negative for MHC class II antigens, as detected by flow cytometry. In contrast to the induction of effector functions, expression of MHC class II antigens stringently required interferon-gamma. IL-1 beta did not induce MHC class I antigen expression. The regulation of ICAM-1 expression in that unstimulated GEC expressed ICAM-1, and this expression was upregulated by IL-1 beta. We conclude that IL-1 beta alters many functions of GEC, and these changes may be involved in the initiation and amplification of glomerular injury.

Interferon-gamma stimulates neopterin release from cultured kidney epithelial cells.

The ability of cultured kidney epithelial cells (KEC) to secrete neopterin, which is a marker of the activation of immune system was studied. In this study interferon gamma (IFN-gamma) was shown to induce neopterin release from KEC in a dose- and time-dependent manner. Many other cytokines and mitogens (IL-1 beta, IL-2, IL-6, LPS and phorbol ester) were tested for their ability to induce neopterin in KEC but all failed to induce a significant release of neopterin from KEC. By itself TNF-alpha induced a release of small amounts of neopterin but strongly potentiated the effect of IFN-gamma in a synergistic manner to induce neopterin secretion. These data indicate that not only monocytes and macrophages which it is well known secrete neopterin, but KEC are responsible also for the high serum or urine level of neopterin observed in patients with kidney allograft rejection or infections episodes. As the amount of neopterin released by KEC is smaller than that secreted by activated macrophages, the contribution of KEC to the overall production of neopterin during certain diseases may be small.

Serum and urinary neopterin as markers in renal transplant patients.

For monitoring neopterin levels, serial serum and urinary samples were obtained from 21 renal transplant patients. In renal transplant patients, serum neopterin levels were significantly higher than in healthy volunteers, even though in a clinically stable state urinary neopterin levels were also higher than in healthy volunteers, but statistically not significantly. In cases with rejection, both serum and urinary neopterin levels were significantly more elevated than in the stable state. In contrast, serum and urinary neopterin levels were not elevated in nephrotoxicity events. These results suggest that serum and urinary neopterin levels might be valuable indicators of acute rejection. Moreover, they could be useful for differentiating acute rejection from nephrotoxicity episode.

Interleukin-8 in chronic renal failure and dialysis patients.

A total of 105 patients participated in this study, including 10 with chronic glomerulonephritis with normal renal function (CGN patients), 36 uraemic patients (CRF patients), 19 continuous ambulatory peritoneal dialysis patients (CAPD) without peritonitis, three CAPD patients with peritonitis, 37 patients undergoing chronic haemodialysis (HD) divided into short-term HD, 15 patients; medium-term HD, 12 patients; and long-term HD, 10 patients. IL-8 and two other proinflammatory cytokines, IL-6 and TNF alpha were tested using a specific immunoassay. IL-8, IL-6, and TNF alpha serum levels were significantly increased in patients with chronic renal failure compared to their levels in normal individuals (P < 0.0001, P < 0.05 and P < 0.0001 respectively). The most pronounced increment in IL-8, IL-6 and TNF alpha serum levels was observed in CAPD patients (P < 0.0001). CAPD patients without peritonitis showed relatively low levels of IL-8 or IL-6 in peritoneal dialysate effluents (PDE), whereas PDE-TNF alpha were not detectable in almost all patients tested. Patients with peritonitis showed very high serum and PDE levels of IL-8, IL-6 and TNF alpha. The clinical recovery from peritonitis was characterized by a rapid fall in IL-8, IL-6 and TNF alpha in serum and dialysate. HD patients showed a significant increase in serum levels of IL-8 and also IL-6 and TNF alpha compared to normal individuals (P < 0.05, P < 0.05 and P < 0.01 respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

Profile of cytokine production in mixed kidney lymphocyte culture.

Kidney cells are an important source of immunoregulatory molecules that regulate cell-to-cell interactions, which is the key step in the generation of the immunoresponse to alloantigens. In this study we identified the cytokines that are produced by both lymphoid cells and kidney cells when coincubated in mixed kidney lymphocyte cultures (MKLC). The capacity of kidney cells to stimulate the proliferation of effector allogeneic lymphocytes was assayed by incubating irradiated kidney cells and lymphocyte. The cytokine secretion profile in MKLC was investigated by incubating monolayers of kidney cells with effector peripheral blood mononuclear cells (PBMC). The culture supernatants were harvested on days 1, 2, 3, 4, 5, 6, 7, and 8 and assayed for IL-1beta, IL-2, IL-6, and TNF alpha using an ELISA. Kidney cells, in comparison to PBMC stimulator cells were poor stimulators of the alloproliferation even when HLA expression was increased by IFN gamma treatment. Compared to lymphocyte or kidney cells incubated alone, MKLC induced a considerable stimulation of cytokine production. This increase in cytokine production was observed essentially for IL-2 and IL-6 (at day 3, a 10-fold increase in IL-2 and a 5-fold increase in IL-6). This study provided evidence that target kidney cells and effector lymphocyte interactions generate a number of cytokines such as IL-1beta, IL-2, IL-6, or TNF alpha. These cytokines are known to modulate allo-proliferation and generation of cytotoxic T lymphocytes (CTL).

Interleukin-8 serum and urine concentrations after kidney transplantation.

We conducted a prospective study of 12 patients undergoing kidney transplantation. In these patients, we monitored interleukin-8 (IL-8) in both serum and urine before and after kidney transplantation. Levels of IL-8 were analyzed by a solid-phase double ligand ELISA method. Three patients with an uneventful recovery after transplantation showed IL-8 serum levels below the detection limit, whereas some small amounts were detected in the urine of these patients. IL-8 serum levels markedly increased with acute graft rejection and infection. Increments in serum and urine preceded clinical complications in all patients. Highest levels were observed in bacterial infection and lowest in acute rejection. Although nonspecific, IL-8 can be considered as an indicator molecule of inflammatory processes occurring during kidney transplantation.

Effect of immunosuppressive agents FK 506 and cyclosporin and steroids on the expression of IL-6 and its receptor by stimulated lymphocytes and monocytes.

The interaction of interleukin-6 (IL-6) with its receptor (IL-6R) is not well understood. In the present study, we investigated the effect of different immunosuppressive agents on the expression of the couple IL-6/IL-6R on cultured lymphocytes and monocytes. IL-6 in culture supernatants from cultured monocytes were analyzed by ELISA. The expression of IL-6R was studied by flow cytometry. Dexamethazone, cyclosporin (CyA), and FK 506 at immunosuppressive concentrations induced a dose-dependent inhibition of IL-6 secretion from adherent monocytes (MO) stimulated with phytohemagglutinin (PHA). Dexamethazone was the most effective agent in inhibiting IL-6 secretion, while the inhibitory effect observed with 1 ng/ml FK 506 was comparable with that obtained with 100 ng/ml CyA. Unstimulated MO strongly expressed IL-6R (80% positive cells). Stimulation of MO with PHA resulted in a significant downregulation of IL-6R expression. Treatment of PHA-stimulated adherent MO with different concentrations of CyA and FK 506 induced a restoration of IL-6R expression. FK 506 was 100 time more effective in restoring IL-6R than CyA. This restoration of IL-6R was incomplete. FK 506, CyA, and steroids may exert their immunosuppressive effect by inhibiting IL-6 secretion and partially restoring MO IL-6R, which may be important in protecting the cell target against IL-6 autocrine stimulation.

[Blood level monitoring of FK506 on kidney transplant recipients].

Blood level monitoring of FK506 was performed on 6 kidney transplant recipients. Reproducibility was evaluated on trough and AUC levels in both whole blood and plasma samples. Reproducibility was almost the same degree on both trough and AUC levels and in both whole blood and plasma samples. Trough and AUC levels in whole blood and AUC in plasma were low at the onset of rejection episodes, whereas those were high at the onset of adverse effects by FK506 such as hyperglycemia, hyperkalemia and neurological symptoms. Frequent trough level monitoring is useful to detect kidney transplant rejection and adverse effects by FK506.

Cytokine-mediated regulation of the surface expression of complement regulatory proteins, CD46(MCP), CD55(DAF), and CD59 on human vascular endothelial cells.

Membrane cofactor protein (MCP), decay-accelerating factor (DAF), and CD59 are complement (C) regulatory proteins that protect human hematopoietic cells from damage induced by autologous complement. Using monoclonal antibodies in both western blotting studies and flow cytometry, we found that endothelial cells (EC) expressed on their surface MCP, DAF, and CD59 molecules. We tested whether stimulation of endothelial cells by known EC activators such as LPS, TNF-alpha, IL-1 beta, and IL-4 regulates the expression of these C regulatory proteins. EC activation was assessed in this study by an up-regulation of ICAM-1 expression. Treatment of EC with various concentrations of TNF-alpha, IL-1 beta, or IL-4, at different incubation times did not increase MCP expression on EC surface membrane. In contrast, the level of DAF was altered during EC activation: LPS or IL-1 beta treatment resulted in a slight increase of DAF expression; the most up-regulatory effect was obtained with IL-4. Up-regulation of surface DAF on EC required prolonged treatment of cells with these agents. The level of CD59 was far greater than that of DAF or MCP on EC, and was slightly up-regulated by TNF-alpha and down-regulated by IL-1 beta. These findings indicate that the levels of C regulatory proteins are regulated in an independent fashion on EC, MCP is not regulated by any cytokine tested, while DAF is an EC activation antigen, although it has a different augmentation profile from ICAM-1 since IL-4 up-regulates DAF but not ICAM-1.