Racemic synthesis and solid phase peptide synthesis application of the chimeric valine/leucine derivative 2-amino-3,3,4-trimethyl-pentanoic acid.

The synthesis of non natural amino acid 2-amino-3,3,4-trimethyl-pentanoic acid (Ipv) ready for solid phase peptide synthesis has been developed. Copper (I) chloride Michael addition, followed by a Curtius rearrangement are the key steps for the lpv synthesis. The racemic valine/leucine chimeric amino acid was then successfully inserted in position 5 of neuropeptide S (NPS) and the diastereomeric mixture separated by reverse phase HPLC. The two diastereomeric NPS derivatives were tested for intracellular calcium mobilization using HEK293 cells stably expressing the mouse NPS receptor where they behaved as partial agonist and pure antagonist.

Chronic periaortitis associated with membranous nephropathy: clues to common pathogenetic mechanisms.

Chronic periaortitis (CP) is a rare disease hallmarked by the presence of a periaortic retroperitoneal fibro-inflammatory tissue which can often cause obstructive uropathy. CP is isolated in most cases but it may also be associated with other sclerosing inflammatory and immune-mediated diseases. We here present the case of a patient who was initially diagnosed as having CP and subsequently developed membranous nephropathy and chronic sclerosing sialoadenitis of the right parotid gland. As these conditions were all characterized by either pronounced infiltration of IgG4-positive plasma cells or marked IgG4 tissue deposition, we hypothesize that they are part of the same disease spectrum, and discuss the immune-mediated pathogenetic mechanisms potentially shared by these conditions. In particular, we consider the role of Th2-mediated immune reactions and of immunogenetic factors such as HLA genotype as common determinants of these disorders.

Tacrolimus Therapeutic Drug Monitoring in Stable Kidney Transplantation and Individuation of CYP3A5 Genotype.

The posology of tacrolimus (TAC) is usually guided by its therapeutic drug monitoring. Some patients reach target concentrations (CTs) quickly, others more slowly. In a retrospective study, 20 kidney transplant recipients were included (mean age, 50.7 ± 14.1 years; weight 64.0 ± 14.2 kg; patients clinically stable for over a year). We studied cytochrome CYP3A5 genotype, in particular CYP3A5 6986A>G, the most important polymorphism related to the metabolism of TAC (wild genotype CYP3A5 *1 genotype, and CYP3A5 *3 variants). One year after transplantation, the CTs were 5.0 to 8.0 ng/mL. The patients were divided into group A (TAC doses < 6.0 mg/d) and group B (TAC doses > 6.0 mg/d). All were tested for the CYP3A5 gene sequence to characterize their polymorphism. Patients with CYP3A5 *1/*1 and *1/*3 were extensive metabolizers, and those with CYP3A5 *3/*3 were poor metabolizers. In group A and group B, the average TAC doses at the time of therapeutic drug monitoring were 3.0 ± 1.4 ng/mL (0.05 ± 0.03 mg/kg) and 12.8 ± 3.7 ng/mL (0.2 ± 0.1 mg/kg), respectively (P < .001). Group A was the poor metabolizers genotype, while in group B, the extensive metabolizers genotype was present. Patients with the CYP3A5 *1/*1 or *1/*3 genotype required 1.5 to 2 times higher doses than patients *3/*3 to reach CT. This genetic test allows clinicians to know, before the kidney transplant, the patient's TAC metabolism pattern and then to optimize the drug exposure.

Interferon-alpha in combination with ribavirin as initial treatment for hepatitis C virus-associated cryoglobulinemic membranoproliferative glomerulonephritis.

Mixed cryoglobulinemia (MC) and glomerulonephritis are the most important extrahepatic manifestations of chronic hepatitis C virus (HCV) infection. In HCV-infected patients with MC, renal involvement worsens the overall prognosis because of a high incidence of infection or cardiovascular disease. The relationship between MC and HCV infection has prompted the use of antiviral therapy. Two patients with chronic HCV infection, type-II MC and membranoproliferative glomerulonephritis (MPGN), presenting as nephrotic syndrome were treated with interferon (IFN)-alpha (3 MU 3 times per week) and ribavirin (15 mg/kg daily) for 6 months. Laboratory tests included measurement of anti-HCV antibodies, HCV RNA, and HCV genotyping, and characterization of circulating cryoglobulins. A pretreatment renal biopsy was performed, and the histopathologic lesions were scored according to the index of disease activity. Viremia and cryoglobulinemia were suppressed in both patients. However, a complete remission of proteinuria was observed in 1 patient only. The evaluation of the renal biopsy specimens revealed a mild MPGN (activity score: 5/24) in the patient with remission of proteinuria and a severe MPGN (activity score: 15/24) in the patient who maintained a nephrotic-range proteinuria. Although a fully satisfactory treatment is not yet available, we feel that a reasonable therapeutic strategy for HCV-infected patients with MC nephritis could be as follows: (1) antiviral treatment alone for patients with a low-grade kidney involvement, and (2) a short-term course of steroids and cytotoxic drugs followed by antiviral therapy for acute exacerbations and/or rapidly progressive GN.

Significance of albuminuria in the follow-up of acute poststreptococcal glomerulonephritis.

The present study was aimed at assessing the diagnostic value of urinary albumin (uA) excretion rate in the long-term follow-up of patients suffering from acute post-streptococcal glomerulonephritis (APSGN). 26 patients, who had presented primarily with nephritic syndrome and showing increased uA without a concomitant rise in total proteinuria (uTP) were followed-up for 131 months on average (range 36-288). At the last check, 14 patients did not show urinary abnormalities, 9 had a persistent increase in uA, 1 increased uTP and 2 renal insufficiency. Urinary and clinical signs of the disease were not seen during observation periods prolonged for 79 months on average (range 20-156) after normalization of uA. No pathological findings were remarked in biopsy specimens obtained in 3 patients when uA was normalized; in contrast, when both uTP and uA (12 cases) or when isolated uA (14 cases) were increased a pattern of diffuse mesangial proliferative glomerulonephritis was constantly observed. These results indicate that the abnormal uA excretion rate during long-term follow-up of APSGN allows to identify a subset of patients with persistent renal disease; conversely, the occurrence of normal uA seems to point to a good diagnostic and prognostic significance.

Mesangioproliferative glomerulonephritis.

UNLABELLED: The clinical and bioptic aspects of 211 cases of primary mesangioproliferative glomerulonephritis with urinary abnormalities lasting mre than 1 year were reviewed. We observed the following clinical syndromes: 1) Persistent proteinuria, isolated or with microhematuria: - a) with latent onset: 39% with hypertension (H); 10% with renal failure (RF); - b) with acute nephritic syndrome at onset: 31% with H; 6% with RF; 2) Recurrent macroscopic hematuria: 26% with H; 10% with RF; 3) Nephrotic syndrome: 70% with H; 29% with RF. The histological lesions, diffuse in all cases, appeared unrelated to the clinical syndromes and/or immunofluorescent patterns. In 65 cases with prevalent mesangial deposits of IgA, 46% showed persistent proteinuria with latent onset, 23% persistent proteinuria with acute nephritic syndrome at onset, 28% recurrent hematuria and 3% nephrotic syndrome. 37% of such patients developed H, 14% RF and 6% remission (R). On the other hand in 65 patients with other deposits the clinical aspects were as follows: persistent proteinuria with latent onset: 46%; persistent proteinuria with acute nephritic syndrome at onset: 31%; recurrent hematuria: 17%; nephrotic syndrome: 6%. 35% of these cases displayed H; 9% RF and 8% R. CONCLUSION: primary mesangioproliferative glomerulonephritis appears to represent a heterogeneous group with only the morphological aspects in common and associated with one of the three above-mentioned clinical syndromes. Immunohistology shows different Ig and/or complement which bear no specific relationship with clinical courses. In particular IgA deposits are present in 50% of the cases and are only related to higher incidence of recurrent hematuria and abnormal IgA serum levels.

Late recurrence of systemic vasculitis after kidney transplantation involving the kidney allograft.

A patient is described in whom a recurrence of vasculitis was observed in the kidney allograft six years after transplantation and one year after withdrawal of corticosteroid therapy. This case shows that systemic vasculitis may recur at any time after transplantation which implies a continuous and careful monitoring of these patients.

Fluorescent in situ hybridisation on tissue sections: a quantitative approach with confocal laser scanning microscopy.

The use of fluorescent detection methods in association with digital microscopy technologies is an innovative approach for tissue localisation of messenger RNA. The success of such methods relies on the tissue preservation, local availability of the probe and on the existence of high resolution tridimensional analysis systems. Cryostatic sections, mild denaturation, short oligonucleotide probes (20mer) and confocal laser scanning microscopy allow the fulfillment of all these conditions avoiding photobleaching and tissue autofluorescence. In this paper, we describe in detail a method for in situ hybridisation set up with digoxigenin-coupled oligonucleotide complementary to beta-actin mRNA as a probe and an anti-hapten fluorescent antibody as second step for detecting specific hybridisation. Fluorescence was analysed by means of a confocal laser scanning microscope (CLSM) that provides images with low out-of-focus blurring also with relatively low numerical aperture (NA) objectives. We propose also an easy method to perform semi-quantitative thresholding analysis which allows to discriminate between background and specific signal.

[Mechanisms of progression of glomerular sclerosis in chronic renal diseases in man]. / Meccanismi di progressione delle sclerosi glomerulari nelle nefropatie croniche dell'uomo.

The available data indicate that in chronic renal failure (CRF) loss of renal function usually progresses at a constant rate toward end-stage renal disease. Although immunological events might be responsible for initiating most glomerular diseases, certain clinical and experimental observations suggest that the rate of progression of these diseases is influenced by several non immunological factors. These factors include systemic hypertension, proteinuria, hyperlipidemia, high protein intake, and probably conditions leading to glomerular hypertrophy. Interventions designed to minimize the potential contribution of these factors to the progression of renal insufficiency may halt or slow the loss of function at early and late stages of CRF.

[Etiopathogenesis of membranous nephropathy: is there a correlation between experimental and human pathology?]. / Eziopatogenesi della nefropatia membranosa: quale correlazione tra la patologia sperimentale ed umana?

The Authors discuss the etiologic, pathogenetic and immunopathologic aspects of Heymann nephritis, in order to compare the numerous acquisitions concerning this nephropathy with the scanty knowledge of human membranous nephropathy, of which it represents the experimental counterpart. This rat disease can be obtained by inoculation of tubular brush border preparations (active form) or of the relevant antibodies (passive form); after an initial hypothesis of glomerular deposition of circulating immune complexes, studies on its pathogenetic mechanisms, instead demonstrated that in situ immunoaggregates, caused by an interaction between circulating antibodies and fixed glomerular antigens, are formed. Recent investigations have led to the identification of a major nephritogenic antigen (gp330), which is a tubular brush border glycoprotein expressed by coated pits located at the glomerular epithelial cell surface. Studies on antigen-antibody interactions at this level have demonstrated that there is a quick redistribution and accumulation of the so-formed immune complexes, and when polyclonal antibodies were utilized, growth of subepithelial electron dense deposits was observed. Although other tubulo-glomerular antigens, which can also be expressed by endothelial cells, play an uncertain role, they seem to favour transmembrane passing of anti-gp330 antibodies. Immune complex formation gives rise to the onset of proteinuria through complement system activation, without leukocyte involvement: in particular a MAC and C9 fraction lytic effect was demonstrated on cultured epithelial cells. In conclusion, studies on Heymann nephritis contribute to our understanding of the etiopathogenetic mechanisms regarding human membranous nephropathy, and emphasize a possible role played by tubular antigens and in situ formed immune complexes.

[Treatment of arterial hypertension in diabetic nephropathy. Certainties and hypotheses]. / Le traitement de l'hypertension artérielle dans la néphropathie diabétique. Certitudes et hypothèses.

Clinical observation has long emphasized the importance of arterial hypertension in the course of diabetic nephropathy and recent studies suggest that hypertension might play a decisive pathogenetic role in the course of the disease, hence the necessity of correcting the hypertension of diabetic patients has by now been universally accepted. There is, however, still some uncertainty concerning the usefulness of acting preventively on so-called microhypertension; in other words, whether early antihypertensive drug treatment can prevent diabetic nephropathy. This paper discusses the criteria to be followed in the choice of antihypertensive medication during diabetic nephropathy giving special attention to pathophysiological considerations. Moreover, it also discusses the effects of antihypertensive drugs currently regarded as first-choice agents, i.e. calcium antagonists and the angiotensin converting enzyme inhibitors, on intrarenal hemodynamics.

[Physiopathology, clinical aspects and prevention of renal insufficiency caused by contrast media]. / Fisiopatologia, clinica e prevenzione dell'insufficienza renale da mezzi di contrasto.

Contrast-media associated nephropathy (CMAN) consists in a sudden impairment of glomerular filtration rate following exposure to radiographic contrast materials. Damage may be limited to an asymptomatic mild increase of blood creatinine, or reach the highest levels of nitrogen retention compatible with acute renal failure. Some preexisting clinical conditions or pathologies may lead to CMAN: not only renal insufficiency, diabetes mellitus, multiple myeloma, congestive heart failure and severe hypertension, but also simple dehydration and a growing series of immunologic diseases are recognized as predisposing condition. The exact mechanism responsible for renal injury is still doubtful but recently animal models have shown substantial ischemic changes that may be added to the traditional presumed pathogenesis of direct tubular toxicity and intra-tubular obstruction. As renal ischemia stimulates both endogenous vasoconstrictor and vasodilator substances, it is now supposed that CMAN acts similarly to non-steroidal anti-inflammatory agents, selectively inhibiting the vasodilatory prostaglandin phase and therefore causing a derangement of the physiologic vasoconstriction/vasodilatation balance of renal circulation. The role of oxygen free radicals to contribute to renal dysfunction is considered. Low osmolality non ionic contrast media when compared to conventional high osmolality ionic contrast media have reduced but not eliminated CMAN. Simple but effective lines of prevention include the previous selection of patients predisposed to CMAN for concomitant pathology, suspension of FANS or any other recognized nephrotoxic substance, the least amount of contrast media compatible with radiologic visualization of the patient's problem, careful hydration of the patient before contrast injection and sustained diuresis afterwards. The usefulness of pre-treatment with Ca-channel blockers or atrial natriuretic factors remains sub judice.

[The treatment of arterial hypertension in diabetes mellitus. Choices and problems]. / Trattamento dell'ipertensione arteriosa in corso di diabete mellito. Scelte e problemi.

Recent studies indicate that arterial hypertension in diabetes mellitus is a paramount pathogenetic step in the evolution and acceleration of diabetic macro- and microangiopathy and in particular in the development of nephropathy and uremia. This paper deals with the clinical problems of antihypertensive treatment in diabetic patients and discusses the antihypertensive repertory with the aim at determining the best drug choice in the individual case. In the light of our present pathophysiologic knowledges of the intrarenal effects of the various classes of antihypertensive drugs the possibility of preventing diabetic nephropathy is discussed.

[Current approach in the prevention and treatment of diabetic nephropathy]. / Attuali orientamenti di prevenzione e trattamento della nefropatia diabetica.

Diabetic renal microangiopathy accounts for enormous morbidity and mortality, particularly in patients who develop diabetes in childhood or early youth; in the last few years its pathogenesis has been therefore extensively studied, aiming to prevent renal complications or at least of slowing down its progression toward uremia. Though not always in accordance with theoretical expectations, the results of clinical trials have nevertheless widened our therapeutic possibilities; in fact, besides the attainment of an optimal metabolic control, other possible interventions include a careful correction of albeit minimal elevations in arterial pressure; the interference with intrarenal hemodynamic parameters; the correction of insulin-independent metabolic pathways, abnormally activated in the diabetic, such as non enzymatic glycation and polyol pathway; the treatment of endothelial and platelet alterations; the improvement of the rheologic properties of blood.

[The physiopathologic bases of the neurotoxicity of phosphorus chelating agents containing soluble aluminum salts in patients with renal insufficiency]. / Le basi fisiopatologiche della neurotossicità dei chelanti del fosforo contenenti sali solubili di alluminio nel paziente con insufficienza renale.

To what extent can damage to the central and peripheral nervous systems be ascribed to chronic aluminum (Al) intoxication taken as a chelating agent for phosphorus, to limit hyperphosphatemia in uremic patients? Since Al is normally eliminated by the renal route, its accumulation in uremia has to be ascribed to a reduced or abolished renal clearance of the metal, which results in preferential toxicity for certain tissues, especially nervous tissue, which shows difficulty in eliminating Al, even after intake has been stopped. This review discusses, on the basis of toxicologic, experimental and clinical data, the possible pathogenic steps of Al neurotoxicity in uremia, considering: the damage to axonal transport in which Al intoxication tends to affect the components of the cytoskeleton, the polymerization phase of the alpha and beta tubulin constituents of neurotubules, and the normal translocation of neurofilaments from the perikaryon to more distal positions of the axon; the abnormalities in the brain pool of adrenergic, cholinergic and GABA neurotransmitters; the increase in permeability and changes in perm-selectivity of the blood-brain-barrier, with further loss of neurotransmitters and with acquisition, from the systemic circulation, of neurotransmitter-like substances such as hormones, monoamines and peptides, which may adversely modulate synaptic and membrane functions; the cerebral energy metabolism and particularly the hexokinase reaction, by Al replacement of the Mg-ion in the Mg-ATP complex, so that phosphorylation of glucose to G6P is blocked; the interaction of Al with calmodulin by displacement of the Ca-ion and subsequent formation of a stable Al-calmodulin complex with a cytotoxic effect due to the increase in the intracellular calcium concentration.(ABSTRACT TRUNCATED AT 250 WORDS)