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Background: Proteinuria is associated with an increased risk of renal failure. In chronic kidney transplant failure it is associated with poorer graft outcome., Materials and Methods: In our Unit 405 renal transplants were performed between April 1992 and December 2001. We analysed 1) the main causes of post-transplant proteinuria and 2) the prognostic significance for graft outcome in patients with a minimum follow-up of 6 months., Results: Early proteinuria was associated with a higher incidence of chronic allograft nephropathy (CAN) and de novo/recurrent nephropathies. Graft outcome was poorer in patients with early persistent proteinuria., Conclusions: Proteinuria after renal transplantation increases the risk of graft failure. We can, therefore, hypothesize that a graft biopsy is the best way to reveal the causes of proteinuria so that therapeutic interventions, which have been shown to reduce proteinuria, can be applied immediately.

The occurrence of pregnancy in young female organ transplant recipients may sustain a high risk for prematurity and low rate of malformations in neonates. Therefore, it is necessary to counsel couples who want a child. In case of pregnancy, strict guidelines must be observed. Continuous exposure to CsA in utero seems to impair T-, B- and NK-cell development and function in neonates. This effect is prolonged throughout the first year of life. In addition, low levels of serum immunoglobulins occur at the same time. This leads to suggest a delayed administration of classical vaccinations (after the first 6 months of life) in view of the potential risks of both sub-optimal immunologic responses, and adverse events after the administration of live, attenuated vaccines in infants born from young female organ transplant recipients.

Over the last few years emerging evidence indicate the involvement of herpes viruses in the pathogenesis of several medical complications in transplanted patients. Herpes viruses are transmitted via inter-human contact and cause a primary infection, which commonly fails to give clinical signs and may persist even for years in a latent state in healthy subjects. In transplanted patients, herpes viruses may be transmitted through the transplanted organ or may be reactivated because of the use of powerful immunosuppressive drugs. Moreover, the persistence of immunosuppression greatly favours the clinical expression and severity of virus infection. Thus, herpes viruses seem to be involved in both acute and chronic deterioration of graft function, in the pathogenesis of post-transplant lymphoproliferative disorders and Kaposi sarcoma, and even in vessel atherosclerosis. This review will focus on relevant clinical aspects of herpes-virus infection, namely cytomegalovirus, EBV, herpes simplex 1 and 2, varicella zoster virus, HHV-6, HHV-7 and HHV-8, in kidney transplanted patients.

Endothelin-1 (ET-1) and nitric oxide (NO) have been suggested to have a focal role in the regulation of placental and fetal growth. Cyclosporine A (CsA) has been shown to strongly modulate ET-1 and NO synthesis and thus has the potential to affect fetal growth and maternal state. Eleven CsA-treated female kidney transplant recipients were recruited. Fourteen healthy pregnant women served as controls. Placental expression of ET-1 and tissue factor (TF) was evaluated by in situ hybridization, and NO synthase (NOS) was evaluated by staining with the reduced form of nicotinamide-adenine dinucleotide phosphate (NADPH)-diaphorase and in situ hybridization. Kidney transplant recipients showed a marked reduction in NADPH-diaphorase staining, as well as endothelial constitutive NOS (ecNOS) messenger RNA, whereas inducible NOS expression was unchanged. Normal placenta showed a strong positive ET-1 signal along the endothelium of uteroplacental arteries within the basal plate, which increased markedly in decidua of transplant recipients. Thus, transplant recipients showed a remarkable alteration in ET-1/ecNOS balance without alteration in fetal growth or maternal renal function. Next, we explored the state of placental endothelial cell activation downstream from vasoactive factors by evaluating TF gene expression. Transplant recipients did not show modification of TF transcript compared with healthy pregnant women. CsA potently affected the placental ET-1/ecNOS vasoactive balance. Nevertheless, newborns from transplant recipient mothers were appropriate for gestational age, and transplant recipients did not show systemic hypertension or impending renal damage. It is suggested that CsA may blunt the activation of endothelial cells and priming of endothelial-derived substances, which possibly lie downstream from the cited vasoactive agents., (Copyright 2002 by the National Kidney Foundation, Inc.)

Background: Delayed graft function (DGF) has been identified as one of the principal correlates of poor graft survival in cadaveric renal transplantation. However, its risk factors and clinical predictors have been poorly elucidated., Methods: We analyzed the risk factors of DGF with a specific emphasis on the role of histological damage of donor kidney. Then, we also studied the impact of DGF, and donor factors affecting DGF, on kidney graft function over the first year after engraftment in 100 consecutive cadaveric renal transplant (Tx) recipients., Results: The organs displaying DGF (n=48) had a significantly higher degree of glomerular sclerosis and tubular atrophy (P<0.01), as well as of interstitial fibrosis and vascular damage (P<0.02) in time-zero biopsies. In patients who received an "ideal" organ for Tx (total histological score < or = 4), DGF showed a strong relationship with Deltaage D-R (70% increase of risk for donors 10 years older than recipients), and with the histological score (odds ratio 1.34). In contrast, donor hypertension was the most relevant variable independently associated with DGF (odds ratio 19.4) in patients receiving a suboptimal organ (histological score >4). Moreover, DGF and donor hypertension adversely affected graft function at 1 year, but only in Tx patients with a histological score >4 in time-zero biopsy. Of note, both patients with and those without DGF showed a very low incidence of biopsy-proven acute rejection (8.5 and 6.8%, respectively) and a rather short cold ischemia time (<16 hr)., Conclusion: Our findings suggest that the quality of the transplanted organ and the occurrence of DGF are strictly related to each other and can influence graft function through apparently nonimmune mechanisms. In addition, long-standing donor hypertension is a strong independent variable affecting both DGF and graft function of suboptimal cadaveric kidneys, at least up to 1 year.

Background: Chronic allograft nephropathy (CAN), the major cause of renal graft failure, frequently displays extensive interstitial fibrin deposition. Little is known in regard to the cause of the altered coagulation/fibrinolysis balance and its relevance in the pathogenesis of CAN. Thrombin, present within the fibrin clots, can interact with a specific receptor, protease-activated receptor 1 (PAR-1), and modulate a variety of cell functions. On the other hand, the derangement of the fibrinolytic system may directly affect extracellular matrix (ECM) degradation., Methods: In the present study, we investigated, by in situ hybridization, PAR-1 gene expression and the mRNA levels for tissue factor and plasminogen activator inhibitor 1 (PAI-1), two key regulatory molecules of coagulation and fibrinolysis, in 16 CAN biopsies and in 10 normal human kidney grafts. The thrombin-induced transforming growth factor beta (TGF-beta) gene and protein expression in proximal tubular cells (PTC) was investigated by Northern blotting and ELISA, respectively., Results: Fibrin deposits, absent in normal grafts, were observed in the interstitial space and arterial wall of CAN. Tissue factor gene expression was not increased either at the vascular or at the interstitial level in CAN. On the contrary, PAI-1 gene expression, barely detectable in control tissue, was strikingly increased in CAN, with a distribution resembling the pattern of fibrin deposition. Note that PAI-1 gene expression was directly correlated with the degree of interstitial fibrosis. In addition, fibrin deposits were strictly associated with a marked increase of PAR-1 gene expression in endothelial cells and PTC. The tubular expression of PAR-1 was significantly higher in Banff grade II-III than in grade I. In vitro, incubation of PTC with thrombin caused a significant up-regulation of TGF-beta gene expression, followed by an increased TGF-beta release into the supernatant. Interestingly, urine from CAN patients contained significantly higher levels of TGF-beta., Conclusions: Fibrin deposits in CAN may result from the increased expression of PAI-1 and the subsequent inhibition of fibrinolysis. The reduced fibrinolysis may cause, in turn, a decreased ECM turnover. Finally, thrombin, preserved in the active form within the fibrin clots, may interact with PAR-1 highly expressed on PTC and induce an up-regulation of ECM deposition in a TGF-beta-dependent manner.

Background: In rodents, CsA has been shown to affect T-cell development, giving rise to an abnormal production of mature T cells and the absence of many T-cell subsets as well as to autoimmunity. Surprisingly, only a few studies investigated the effect of the immunosuppressive drug on the immune system of the human fetus., Methods: We examined six infants born to female kidney transplant recipients who had received cyclosporine and methylprednisolone throughout their pregnancies. Peripheral blood was obtained 1 day and 2, 4, 6, and 12 months after birth, and two-color flow cytometric immunophenotyping of lymphocytes was performed., Results: Total T cells, as well as CD4+ and CD8+ T cells, were low at birth, but normalized thereafter. Among T-cell activation markers, the expression of CD25, the alpha chain of the interleukin-2 receptor, was below the normal range or low range throughout the study period, and HLA-DR expression was extremely low at birth and failed to increase up to 12 months. The number of total B cells was lower than normal at birth, but steeply increased over time. In contrast, B-cell subset bearing CD5 antigen was severely depleted throughout the first year of life. Total IgG concentration was significantly lower than in controls at 2 months, mainly because of subnormal levels of IgG1 and IgG3 subclasses, which remained in the low range up to 6 months. Finally, infants showed normal numbers of true natural killer (NK) cells (CD3-CD16+CD56+), whereas the expression of CD57 antigen, defining non-MHC-restricted cytotoxic lymphocytes, was barely detectable at birth and failed to increase over time, in both CD8+ and CD8- subsets. Of note, none of the infants had clinical evidence of an immunodeficient state., Conclusions: continuous exposure to CsA in utero seemingly impairs T-, B-, and NK-cell development and/or maturation, and most of its effects are still apparent at 1 year, which might suggest that conventional vaccinations should be delayed in these infants.

Hyperlipoproteinemia has been reported to frequently occur in kidney transplanted patients, thus possibly explaining, at least in part, the increased incidence of cardiovascular disease in this population. To evaluate the impact of renal transplantation (Tx), and related immunosuppressive therapy, on plasma lipoprotein and Lp(a) profile, we selected a cohort of kidney transplanted patients (36 M/14 F; age 33.8 + 12.0 yr, range 13-62) lacking significant causes of hyperlipidemia. All patients received a triple immunosuppressive regimen and showed a stable renal function after Tx (plasma creatinine: 1.36 +/- 0.35 mg/dL). One year after Tx, we found a significant increase of total cholesterol (TC), LDL, HDL, ApoB and ApoA-I (p < 0.005), while plasma triglyceride levels remained unmodified. Lp(a) plasma levels after Tx were within the normal range and displayed a significant inverse relationship with apo(a) size. Noteworthy, LDL/HDL ratio and ApoB/ ApoA-I ratio in kidney transplanted patients were almost superimposable with those of normal controls. Specifically, LDL/HDL ratio significantly decreased in 64% of patients after Tx, due to a prevalent increase of HDL, and was associated with a moderate amelioration of plasma TG. In a multiple linear regression model, post-Tx HDL level was significantly related to recipient's age, gender, BMI and cyclosporine (CyA) trough levels (Adj-R2 = 0.35, p = 0.0002), with gender and CyA trough levels being the better predictors of HDL. In conclusion, immunosuppressive regimens, in themselves, do not appear to significantly increase the atherogenic risk related to lipoproteins. Rather, other factors can affect the lipoprotein profile and its vascular effects in renal transplant recipients.

Cyclosporin (CsA) is widely used in the treatment of renal disease and transplantation, which are often complicated by alterations of lipid metabolism. Both chronic administration of CsA and hyperlipidaemia have been shown to evoke an early macrophage influx and have progressively led to glomerular and interstitial sclerosis. MCP-1 is the major monocyte chemoattractant secreted by stimulated mesangial cells and TGF-beta 1 is a key mediator of fibrogenesis in chronic progressive renal fibrosis. Thus, the combined effect of CsA and low-density lipoprotein (LDL) on the gene and protein expression of MCP-1 and TGF-beta 1 in cultured human mesangial cells (HMC) was explored. Both agents induced an early and persistent increase of MCP-1 and TGF-beta 1 mRNA levels and protein release. The simultaneous addition of CsA and LDL did not display any additive effect on target gene expression, but it caused a synergistic effect on MCP-1 and TGF-beta 1 protein secretion into culture medium. On the other hand, CsA and LDL had different effects on cell proliferation: the latter increased DNA synthesis, whereas CsA inhibited both spontaneous and mitogen-stimulated mesangial cell growth. The study concludes that CsA and LDL display an additive effect on TGF-beta 1 and MCP-1 synthesis and release by HMC, thus possibly co-operating to induce an early macrophage influx and the subsequent mesangial expansion and increased extracellular matrix deposition. However, in contrast they seem to modulate HMC proliferation differently, which is a further critical event intimately involved in the development of glomerulosclerosis.

One of the major concerns in space medicine is post-mission postural hypotension. Since the beginning, most of the studies have been aimed at the identification of mechanical and neuroautonomic mechanisms within the cardiovascular system potentially involved in this phenomenon. Some gender differences in the cardiovascular reactivity to changing posture have been described so far, which still need better clarification. Moreover, the neuroendocrine contribution to postflight readaptation deserves more indepth studies. Sudden passive head-up tilting is considered to be a good simulation method for hemodynamic changes occurring during transition from 0xg to 1xg. The aim of this study was to verify whether cardiovascular or neuroendocrine changes occurring after sudden passive +70 degrees head-up tilt (HUT) became already evident within 10 min and were similar in healthy young-adult men and women.

Background: Recent data indicate that factors other than erythropoietin (EPO), such as insulin-like growth factor 1 (IGF-1), can promote erythropoiesis in vitro and correct the anemia of chronic renal failure in vivo. IGF-1 is produced by the liver under growth hormone control, as well as by other sources, including the kidney. The erythropoietic role of growth factors and cytokines and their possible modulation by angiotensin-converting enzyme inhibitors (ACEI) has never been explored., Methods: This study evaluated the serum levels of EPO, IGF-1, interleukin (IL)-2, IL-3, and granulocyte macrophage-colony-stimulating factor in 40 kidney transplanted patients with or without posttransplant erythrocytosis (PTE) and in 10 living kidney donors. Then, the effect of ACEI therapy on the above pattern was examined in patients with PTE., Results: EPO and IGF-1 serum levels were significantly higher in patients with PTE than in patients without PTE and in living kidney donor subjects. ACEI therapy significantly reduced hematocrit (Hct) as well as circulating IGF-1 and EPO levels. Of note, the decrease in IGF-1 was prominent mainly in those patients whose EPO levels were not significantly modified by ACEI therapy. In all of the patients Hct levels displayed a direct relationship with circulating IGF-1 levels, but not with EPO concentration. Growth hormone did not significantly differ among the groups examined, whereas it steeply increased under ACEI. Finally, no significant difference in IL-2, IL-3, and granulocyte macrophage-colony-stimulating factor serum levels was detected., Conclusions: IGF-1 seems to play a role in the ACEI-related decrease of Hct in patients with PTE, chiefly in patients without any modification of EPO serum levels.

High glucose concentration has been shown to induce the overexpression of transforming growth factor (TGF)-beta 1 mRNA and protein in different cell types, including murine mesangial cells, thus possibly accounting for the expansion of mesangial extracellular matrix observed in diabetic glomerulopathy. In the present study, we evaluated platelet-derived growth factor (PDGF) B-chain and PDGF-beta receptor gene expression in human mesangial cells (HMCs) exposed to different concentrations of glucose and then sought a possible relationship between a PDGF loop and the modulation of TGF-beta 1 expression. HMC [3H]thymidine incorporation was upregulated by 30 mmol/L glucose (HG) up to 24 hours, whereas it was significantly inhibited at later time points. Neutralizing antibodies to PDGF BB abolished the biphasic response to HG, whereas anti-TGF-beta antibodies reversed only the late inhibitory effect of hyperglycemic medium. HG induced an early and persistent increase of PDGF B-chain gene expression, as evaluated by reverse transcriptase polymerase chain reaction, whereas PDGF-beta receptor mRNA increased by twofold after 6 hours, thereafter declining at levels 70% lower than in controls after 24 hours. 125I-Labeled PDGF BB binding studies in HMCs exposed to HG for 24 hours confirmed the decrease of PDGF-beta receptor expression. TGF-beta 1-specific transcripts showed 43 and 78% increases after 24 and 48 hours of incubation in HG, respectively, which was markedly diminished by anti-PDGF BB neutralizing antibodies or suramin. We conclude that HG induces an early activation of a PDGF loop that, in turn, causes an increase of TGF-beta 1 gene expression, thus modulating both HMC proliferation and mesangial matrix production.

The kidney is one of the major sites of EGF production and there it seems to play several biological functions, such as modulation of cell growth, renal repair following injury, regulation of cellular metabolism and glomerular haemodinamics. The present study was first aimed at localizing EGF and its receptor (R) in normal human kidney by immunohistochemical and in situ hybridization techniques. Then, the distribution of the growth factor and its R was explored in biopsy specimens from eight patients with acute tubulointerstitial damage. In the normal human kidney, both EGF immunoreactivity and EGF mRNA were localized in tubular profiles corresponding to Henle's loop and, although to a lesser intensity, to distal convoluted tubule. EGF immunostaining was remarkable mainly at the apical surface of tubular cells. EGF-R protein expression was detected in glomerular endothelial cells, in peritubular capillaries and arteriolar walls, as well as along the thick ascending limb of Henle's lop and distal convoluted tubule, where it colocalized with Tamm-Horsfall protein. Immunohistochemical analysis of tubular profiles revealed that EGF-R was located especially along the basolateral membrane of tubular cells and within the basal part of cytoplasm. Endogenous alkaline phosphatase and CHIP28 positive tubules did not show any signal for EGF and its receptor. Kidneys with acute tubulointerstitial injury exhibited a dramatic decrease of EGF expression, whereas EGF-R showed only minor modifications. Interestingly, EGF-R was localized to both apical and antiluminal membranes of positive tubular cells. It is concluded that EGF-EGF receptor loop may be relevant in the pathogenesis of acute tubulointerstitial damage and recovery from tubular injury, while its role in the physiological renewal of the urothelium remains speculative.

Cultured human mesangial cells (HMC) derived from normal kidneys have been shown to synthesize tissue-type plasminogen activator (t-PA) and excess amounts of PA inhibitor type 1 (PAI-1). Conflicting results have been obtained concerning the production of urokinase-type PA (u-PA) and efforts to show PA inhibitor 2 (PAI-2) met with failure. We evaluated the fibrinolytic profile of cultured HMC lines obtained from 12 patients with renal carcinoma and one cadaveric kidney donor. Subconfluent cells (third passage) were incubated overnight in serum-free medium. t-PA, u-PA, PAI-1 and PAI-2 antigens were assayed by ELISA methods and PA and PAI activities by amidolytic methods both in conditioned medium (CM) and cell extracts (CE). Besides PAI-1, PAI-2 antigen was detected in all but one HMC lines. At variance with the former, which was largely released in the culture medium, PAI-2 was mainly cell-associated. t-PA antigen was found in all but two cell lines while u-PA antigen was detected in relatively high concentrations in 8 cell lines. PA activity, identified as u-PA by functional and immunological criteria, was measured in CM of six of the eight u-PA producing cell lines, whereas PAI activity was undetectable or very low in CM of all cell lines, suggesting that PAI-1 was largely inactive. Functional assays of cell extracts demonstrated the presence of PA activity, again identified as u-PA, only in samples (five lines) containing u-PA antigen in excess over PAI-2. PAI activity was found instead in the extracts in which the inhibitor was higher than the activator (six lines) and was identified as PAI-2, as it inhibited u-PA but not single-chain t-PA and was neutralized by a polyclonal anti-PAI-2 antibody. The heterogeneous fibrinolytic pattern of HMC lines was confirmed by mRNA analysis of three representative lines. Results were similar when HMC lines at passage five were used, except that the u-PA content was significantly reduced both in CM and CE. These findings indicate that the fibrinolytic profile of cultured HMC is more complex than previously reported. The production of large amounts of PAI-2 may represent an additional control mechanism of proteinase activity.

Mesangial cell (MC) proliferation, a histopathologic feature common to many human glomerular diseases, is regulated by several growth factors through their binding to specific cell surface receptors. Platelet-derived growth factor (PDGF) is a peptide exerting a potent mitogenic activity on MC. Recently, an increased expression of both PDGF protein and its receptor has been localized in the mesangial areas of several experimental as well as human proliferative glomerulonephritides (GN). Thus, it may be postulated that the inhibition of PDGF action could prevent MC proliferation during mesangial proliferative GN. Trapidil, an antiplatelet drug, has been shown to inhibit the growth of several cell types both in vitro and in vivo. The present study was aimed at evaluating the effect of Trapidil on human MC in vitro. The addition of 100 to 400 micrograms/ml Trapidil significantly reduced cell proliferation induced by different growth factors (FCS, PDGF-BB, bFGF, EGF), the highest inhibitory effect being on PDGF-BB stimulated MC growth. The effect of the drug was dose-dependent and seemingly specific: aspirin was devoid of any anti-proliferative action, while dypiridamole proved to be toxic. Receptor binding experiments showed that Trapidil competitively inhibited 125I-PDGF-BB binding to its cell surface receptors, without inducing receptor internalization, at least after short-term (2 hr) incubation. In contrast, long-term (48 hr) exposure to 400 micrograms/ml Trapidil caused a sharp increase of PDGF-BB binding. Similar effects on cell proliferation and 125I-PDGF-BB binding were observed when NIH-3T3 fibroblasts were exposed to the test substance.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of metformin on insulin binding and insulin action in the presence of anti-insulin receptor antibodies was investigated in a case of type B extreme insulin resistance. Oral administration of metformin (1500 mg/d) for 10 days significantly decreased plasma blood glucose and insulin levels and enhanced the hypoglycemic response to exogenous insulin. In vitro preincubation of normal erythrocytes with insulin receptor antibody from the patient plus 4 x 10(-5) mol/l metformin markedly enhanced insulin binding to receptors, compared to cells incubated with antibody alone. This effect was apparent after 2 h, was maximal after 4 h and did not change up to 24 h. Closely similar results were found when human adipocytes were studied. Analysis of binding data confirmed the increase in both receptor number and affinity. One hour exposure of control adipocytes to metformin enhanced basal lipogenesis by more than 30%. Acute exposure of fat cells to the patient's receptor antibodies resulted in a stimulation of glucose transport and a state of severe insulin resistance. The addition of metformin to antibody in preincubation buffer strongly enhanced basal glucose incorporation into lipids, but did not prevent insulin unresponsiveness. It is suggested that metformin increases, possibly through a change in the spatial conformation of insulin receptor within the plasma membrane, the availability of pre-existing receptors to insulin binding and/or decreases the availability of specific epitopes to antibody anchoring. Further, in the model of insulin resistance described here, metformin enhanced the basal rate of glucose transport through a direct insulin-mimicking activity and/or a potentiation of the sensitivity of glucose transport to the antibody.

We studied a patient with systemic lupus erythematosus and type B insulin resistance, who progressed from extreme insulin resistance to fasting hypoglycemia. The plasma insulin level was 63.3 +/- 20.9 pmol/L in the fasting state and rose above 1440 pmol/L postprandially. Intravenous administration of human insulin caused almost no decline in plasma glucose. Therefore, it was concluded that the patient was still resistant to insulin and that plasma insulin did not play a crucial role in the development of hypoglycemia. Immunoglobulin G from this patient did not inhibit insulin binding to the insulin receptor; rather, it enhanced [125I]insulin binding in both the immunoprecipitate and the in vitro binding assay to intact cells. Antiinsulin receptor antibodies strongly inhibited insulin internalization in human adipocytes, slowed down the dissociation of [125I]insulin from receptors and failed to induce down-regulation of surface insulin receptors in both the presence and absence of insulin. Finally, autoantibodies mimicked the insulin stimulatory effect on human fat cell lipogenesis even after long term exposure, but inhibited the metabolic potency of insulin when added simultaneously with the natural ligand. We conclude that antiinsulin receptor antibodies induce fasting hypoglycemia, through their continuous receptor stimulatory action, and insulin resistance, possibly by a conformational perturbation of the receptor protein, which, in turn, uncouples insulin receptor binding from receptor function.

The effect of D-(-)-beta-hydroxybutyrate, at concentrations commonly achieved during ketoacidosis in humans (10 mmol/l), on human fat cell lipolysis in vitro was the aim of this study. The basal lipolysis was not modified and beta-hydroxybutyrate did not affect forskolin- or dibutyryl-cAMP-stimulated glycerol release, whereas it markedly inhibited isoproterenol-stimulated lipolysis. In membranes of intact adipocytes exposed to D-(-)-beta-hydroxybutyrate for 1 h, we found a decrease in beta-adrenoceptor affinity in saturation experiments and a shift to the right of the isoproterenol-mediated radioligand [( 125I]-cyanopindolol) displacement curve. These findings suggest that beta-hydroxybutyrate inhibits catecholamine-stimulated lipolysis by decreasing beta-adrenoceptor affinity. No effect of beta-hydroxybutyrate was found on beta-adrenoceptor binding of intact mononuclear cells of peripheral blood. In conclusion, the beta-adrenoceptor affinity lowering effect of beta-hydroxybutyrate is seemingly specific to human fat cells and might represent a feed-back mechanism that prevents an uncontrolled breakdown of triglycerides and indirectly regulates its own production rate.

381 glucose intake normal curves were studied according to the Diabetes Data Group new classification in healthy persons between 10 and 80 years in order to assess the influence of the age upon the normal glucose tolerance. Such an influence, which was evident in all the subjects, turned out to be more important in women with respect to me. In fact, males showed an increase, per decade, of about 1 mg/dl in fasting glycemic levels, of about 6 mg/dl at 60', of 4 mg/dl at 120', while in females there was an increase of about 2 mg/dl in fasting glicemic values, of about 6 mg/dl at 60' and of about 5 mg/dl at 120'. No meaningful correlation between age and insulinemic values was found at all considered points, either in males or in females. The reasons of the decreased glucose tolerance with aging and of its different behavior in the two sexes are discussed.

The influence of lactate on human adipocytes lipolysis and the possible relationship between lactate-induced metabolic effects and beta-adrenoceptor binding sites were investigated. beta-sites were identified in membranes with (125I)-cyanopindolol and in intact cells with (125I)-cyanopindolol and (3H)-CGP 12177. Lactate reduced isoproterenol-induced lipolysis in a dose-response fashion and such inhibition became significant only at 16 mmol/l lactate. Exposure of human fat cells to 16 mmol/l lactate significantly reduced beta-adrenoceptors density on crude membranes. When the binding assay was performed on intact cells using (125I)-cyanopindolol at 37 degrees C, the radioligand identified the same number of receptors, regardless of the presence of lactate in the preincubation medium. When (3H)-CGP 12177 was used, it bound to about 35% less receptors in lactate pre-treated cells than in control. Seemingly, at 37 degrees C, because of its lipophilicity, (125I)-cyanopindolol can cross the plasma membrane and bind to intracellular sites whereas, (3H)-CGP 1277, due to its hydrophilicity, identifies surface receptors only. Thus, the present in vitro study provides evidence that high levels of lactate, similar to the concentrations usually achieved in overt lactic acidosis, are able per se to inhibit human lipolysis and to redistribute beta-adrenoceptors from cell surface to a domain not accessible to hydrophilic ligands.

In the present study we evaluated somatomedin-C (Sm-C) plasma levels in diabetic patients, with and without retinopathy. One hundred and thirty four diabetic patients (65 type I and 69 type II) and 90 controls, strictly matched for age and sex, were enrolled in the study. Ophthalmoscopy and fluorescein angiography allowed to distinguish: 49 patients without retinopathy, 45 patients with background retinopathy, and 40 with proliferative retinopathy. Growth hormone (GH) and Sm-C plasma levels were measured using a pool of 20-24 blood samples over 24h. Sm-C levels in type I (0.62 +/- 0.11 U/ml) and type II (0.56 +/- 0.09 U/ml) patients were significantly decreased (p less than 0.01) when compared to controls (0.89 +/- 0.30 U/ml). The mean daily secretion of GH was significantly (p less than 0.01) greater in diabetic patients (7.8 +/- 2.6 ng/ml) than in controls (4.1 +/- 1.5 ng/ml), but no correlation was found between Sm-C and GH (r = 0.15; p = n.s.). Our findings did not show any correlation between Sm-C plasma levels and either the existence of retinopathy, regardless of the degree of microvascular damage, or duration of the disease, or degree of metabolic control, as evaluated by HbA1c.

Fibronectin is a high molecular weight alpha-2-glycoprotein. Its peculiar role in the structure of connective tissue, together with its wide involvement in coagulative dynamics, justified the increasing interest for fibronectin in the pathogenesis of diabetic disease and its vascular sequelae. In the present work, we evaluated the levels of plasma fibronectin (PF) in diabetics with and without retinopathy, and studied the possible correlation between the glycoprotein and some hormonal and metabolic parameters, expression of glycometabolic balance. We examined 26 type I and 24 type II diabetics, further divided into retinopathics and not retinopathics, and 43 normal subjects. We did not find any significant difference in PF levels either between normals and diabetics, or between type I and type II patients, or between retinopathics and not retinopathics. PF was significantly correlated to age, both in normals and in diabetics. Diabetic patients showed a significant positive correlation of PF to total cholesterol (r = 0.56; p less than 0.05) and triglycerides (r = 0.36; p less than 0.05). This seems to suggest, although indirectly, the existence of a relationship between the levels of PF and the degree of large vessel involvement. No significant correlation was found with HbA1c, beta-OH, AcAc, lactate, pyruvate, C-peptide, total and free insulin or GH. We further indicated an inverse correlation between PF and plasma glucagon (IRG). Very low levels of PF are commonly associated with high IRG plasma values during acute energy deprivation such as prolonged fasting and ketoacidotic coma. Therefore, PF levels might represent an index of latent to overt energy depletion.

Catecholamines acutely exert a pronounced insulin-antagonistic effect, which is mediated by beta-adrenergic receptors stimulation. Nevertheless, several patients with pheochromocytoma fail to exhibit an overt diabetic syndrome, in spite of steadily elevated plasma levels of catecholamines. This prompted us to investigate a 16 years old male patient, bearing an extra-adrenal pheochromocytoma, who displayed a slightly impaired glucose tolerance to oral glucose tolerance test, whereas fasting and post-prandial blood glucose, as well as glycaemic response to intravenous glucagon, were in the normal range. Peripheral insulin sensitivity, as evaluated by intravenous insulin tolerance test, was slightly decreased. Supine norepinephrine plasma levels were steadily upon 9 ng/ml; plasma insulin, both fasting and post-prandial, was within the normal range. beta-adrenergic receptors density of peripheral mononuclear cells was strongly reduced when compared to controls (0.97 +/- 0.08 vs 2.82 +/- 0.37 fmol/10(6) cells), without any concomitant change of affinity. Insulin binding to circulating monocytes was reduced as well (2.38 +/- 0.27 vs 5.1 +/- 0.4%/10(7) monocytes); insulin receptor affinity was quite normal (1.7 ng/ml) and total receptor number was 9,200 sites/cell. In desensitization experiments, 1 microM isoproterenol caused only a 20% decrease of beta-adrenergic receptors density in the patient's cells (70% decrease in controls). Six months after surgery, all the above modifications of receptor binding, as well as the mild glucose intolerance, were almost completely reversed. Thus, high levels of norepinephrine were able to induce a decrease of both beta-adrenoceptor and insulin receptor binding, together with a marked reduction of in vitro agonist-induced redistribution of beta-adrenergic receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Optimal and early control of recent onset, type I diabetes by intensive insulin therapy has been reported to allow insulin withdrawal in about two thirds of subjects treated. We used continuous s.c. insulin infusion (CSII) in the attempt to induce a temporary remission of insulin dependence in 18 newly diagnosed young adult diabetics. After 10 days of optimized glycometabolic control, insulin infusion was stopped and patients were switched to glibenclamide (15 mg/die) plus metformin (1 g/die). Diabetics were considered in remission of insulin dependence when their metabolic control fulfilled the following criteria for at least 3 months: absence of glycosuria, pre- and post-prandial blood glucose less than or equal to 120 and 180 mg/dl, respectively, HbA1c less than or equal to 7%. Insulin therapy could be discontinued for periods of over three months in 11 subjects (61%) and for as long as 18 months in one case. Insulin requirement during CSII was slightly higher in nonremitters (NR) than in remitters (R): 0.36-0.64 vs 0.26-0.41 U/kg/die. After 24 months from CSII, R still showed lower insulin requirement (0.35-0.42 U/kg/die) than NR (0.55-0.75 U/kg/die). Further, the role of some hormonal and immunologic factors was investigated. Plasma C-peptide and glucagon were measured, fasting and 2h after each meal, both on admission and immediately after CSII, when patients were switched to oral therapy. No difference in hormone levels could be detected on admission, whereas, after CSII, mean post-prandial increase of C-peptide over basal was significantly higher in R than in NR (1.18 +/- 0.37 vs 0.22 +/- 0.16 ng/ml, p less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)