Selection of sucrose-dependent Escherichia coli to obtain envelope mutants and fragile cultures.

Mutants isolated as sucrose-dependent include many with apparent envelope defects, which fact often leads to filamentous growth or lysis in the absence of sucrose. One mutant can grow exponentially in 4 percent sucrose, but is very fragile: it releases all of its RNA and soluble protein when treated with 0.5 percent sodium deoxycholate. These characteristics permit the study of unstable structures and rapid processes in actively growing cells.

Rates of synthesis and degradation of ribosomal ribonucleic acid during differentiation of Dictyostelium discoideum.

Synthesis of ribosomes and ribosomal ribonucleic acid (RNA) continued during differentiation of Dictyostelium discoideum concurrently with extensive turnover of ribosomes synthesized during both growth and developmental stages. We show here that the rate of synthesis of 26S and 17S ribosomal RNA during differentiation was less than 15% of that in growing cells, and by the time of sorocarp formation only about 25% of the cellular ribosomes had been synthesized during differentiation. Ribosomes synthesized during growth and differentiation were utilized in messenger RNA translation to the same extent; about 50% of each class were on polyribosomes. Ribosome degradation is apparently an all-or-nothing process, since virtually all 80S monosomes present in developing cells could be incorporated into polysomes when growth conditions were restored. By several criteria, ribosomes synthesized during growth and differentiation were functionally indistinguishable. Our data, together with previously published information on changes in the messenger RNA population during differentiation, indicate that synthesis of new ribosomes is not necessary for translation of developmentally regulated messenger RNA. We also establish that the overall rate of messenger RNA synthesis during differentiation is less than 15% of that in growing cells.

Different mRNAs have different nuclear transit times in Dictyostelium discoideum aggregates.

Nuclear processing of mRNA precursors in differentiating multicellular Dictyostelium discoideum aggregates is markedly slower than in growing amoebae. Thus, we have been able to determine the time of nuclear processing of individual mRNA species in postaggregating cells by following the incorporation of 32PO4 into nuclear and cytoplasmic RNA complementary to cloned cDNAs. Precursors of mRNAs synthesized during both growth and differentiation remain in the nucleus for about 25 to 60 min. By contrast, typical mRNAs which are synthesized only by postaggregative cells have nuclear processing times between 50 and 100 min. Depending on the particular mRNA, between 20 and 60% of nuclear transcripts are converted into cytoplasmic mRNA. A third class of mRNAs are transcribed from a set of repetitive DNA segments and are expressed predominantly during differentiation. Nuclear precursors of these mRNAs are extensively degraded within the nucleus or very rapidly after transport to the cytoplasm. Those sequences that are stable in the cytoplasm exit from the nucleus only after a lag of over 2 h. Thus, mRNAs encoded by different genes that are subject to different types of developmental controls display different times of transit to the cytoplasm and different efficiencies of nuclear processing. Differential nuclear processing may contribute to the regulation of the level of individual cytoplasmic mRNAs.

Cell type specificity and mechanism of control of a gene may be reverted in different strains of Dictyostelium discoideum.

Twelve genes which are expressed exclusively in pre-spore cells of Dictyostelium strain AX3 are expressed exclusively in pre-stalk cells of strain AX2. One gene has the opposite behavior: it is expressed in pre-stalk cells in AX3 and in pre-spore cells in AX2. The change in cell type specificity involves a change in the mechanism of control of gene expression. When they are expressed in pre-stalk cells, genes are controlled at the level of transcription, whilst in pre-spore cells, they are controlled at the level of mRNA stability. Genes expressed in pre-stalk cells in strain AX2, fused with an AX2 pre-spore specific promoter, become regulated at the level of mRNA stability. These findings indicate that at least a group of pre-stalk mRNAs possess the cis-destabilizing element typical of pre-spore mRNAs, though they are not destabilized in disaggregated cells. This is due to the fact that ribosomal protein S6, phosphorylation of which is responsible for controlling the stability of pre-spore mRNAs, is not dephosphorylated in disaggregated pre-stalk cells. These cells lack an S6 phosphatase activity which has been purified from disaggregated pre-spore cells.

Control of mRNA stability during development of Dictyostelium discoideum.

A large group of mRNA species (which are mainly pre-spore specific) accumulate only after the formation of multicellular aggregates. They are transcribed at a constant rate from the beginning of development and their accumulation is controlled by a 10-20-fold increase in their stability. This mRNA stabilization is dependent upon multicellularity. When aggregates are dispersed, the mRNAs are destabilized; if cells are allowed to reaggregate, the destabilization is reversed. Destabilization is not due to a selective exclusion of mRNA from polyribosomes, but is a primary control event. It does not require synthesis of new RNA or protein, but it may require an interaction between ribosome and the 5'-end of mRNA molecules.

Beta-2-microglobulin serum profiles in different settings of mass transport and fluid pyrogen content.

Different beta 2-microglobulin (beta 2m) serum profiles have been related to dialytic membranes, mass transport and/or patient immune stimulation. Eight patients were followed by cycles of four sessions: hemodialysis (HD), hemodiafiltration (HDF), acetate-free HDF (AFH), hemofiltration (HF) by filters on synthetic membranes (polysulphone = 4; methylmethacrylate = 4); pre- (A) and post- (B) measurements in the fourth session, and at the start of the next one (C), beta 2m lipopolysaccharide content of the fluids (LPS), and monocytes in vitro and spontaneous production of interleukins (IL); IL-1-IL-6 and tumor necrosis factor (TNF) were measured. In HD, beta 2m (mg/liter), corrected for ECV distribution, did not change (A = 36.5 +/- 10, B = 37 +/- 9, C = 36.4 +/- 9.7). In HDF, lower basal beta 2m (P < 0.001; A = 26.5 +/- 9) still decreased (B = 9.13 +/- 6.2), boosting subsequently to C = 21.6 +/- 14, as in AFH (A = 24.5 +/- 7, B = 11.2 +/- 2, C = 25.3 +/- 9) and in HF (A = 26.6 +/- 7; B = 8.5 +/- 4; C = 25.6 +/- 11). LPS (EU/ml) decreased (P < 0.001) from HD fluids (0.41 +/- 0.1) to HDF (0.28 +/- 0.1), AFH (0.15 +/- 0.1) and HF (0.04 +/- 0.05) but IL-1 and IL-6 were found in greater concentrations in HDF and AFH versus HD and HF, probably due to back-filtration. Beta 2m in different modes of dialytic treatments seem better correlated with the amount of convective transport rather than with the selected markers of immune stimulation.

Impaired fibrinolysis in uremia: partial and variable correction by four different dialysis regimes.

In addition to a hemorrhagic diathesis, uremia is accompanied by a clotting tendency, caused by a marked fall in fibrinolytic capacity. Measurement of lysis time of whole blood diluted with phosphate and acetate buffers and of euglobulin lysis times showed that accumulation of inhibitors is primarily responsible. These probably belong to the class of small molecules abnormally retained in uremia. Hemodialysis (HD) offers the best method of correction, mainly because of better elimination of these inhibitors. In contrast, hemofiltration (HF) and, particularly, intermittent peritoneal dialysis (IPD) are much less effective. In IPD, protein loss via the peritoneum is also responsible for a loss of fibrinolytic activators, so that fibrinolysis becomes even poorer, exposing the patient to an increased risk of vascular complications.

Heparin transfer across the rabbit peritoneal membrane.

Variable quantities of heparin have been proposed to avoid intraperitoneal clotting during peritoneal dialysis without the risk of systemic effects, because heparin is presumed to be incapable of passing through the peritoneal membrane. This study set out to verify this assumption by using labeled heparin in experimental dialysis in 7 New Zealand white rabbits. Heparin was labeled with 99mTc. Labeling quality, assessed by two chromatographic checks, showed less than 5% of free pertechnetate. Chromatographic determinations showed more than 95 and 80% of labeled heparin in inflow and outflow dialysates and in blood samples respectively. Following sodium thiopental anesthesia, animals underwent three protocols: a single 15 min cycle of time diffusion with heparin 500 U/l (A), 6 successive 15 min cycles with heparin 500 U/l (B), and a single 3 h cycle with heparin 2,500 U/l (C). Labeled heparin was found in blood organs and urine in variable percentages. The total amount of recovered radioactivity ranged from 1.5% (A) to 20% (C) of that introduced. It may be concluded that heparin passes through the peritoneum according to some law dependent on the amount used and the diffusion time.

Prevention and treatment of aluminum overload in uremic patients: long-term results.

The authors evaluate the efficacy of a protocol of prevention and treatment of aluminum (Al) overload in RDT patients during a 7-year period (from 1981, 164 patients, to 1987, 161 patients). Al in dialysate solutions was always less than 25 micrograms/l. Baseline Al levels greater than 100 micrograms/l were found in 22% of patients in 1981 but in none in 1987, while the percentage of values less than 60 micrograms/l increased from 55 to 91%. DFO tests were positive in 54% and 7% of cases in 1981 and 1987, respectively. A clinical diagnosis of Al intoxication was performed in 6 patients in 1981, and no further cases were diagnosed later. DFO treatment (50 mg/kg once a week) was employed preventively in 31 patients owing to positive DFO-tests, and in the 6 Al-intoxicated patients therapeutically. In the former patients none developed clinical intoxication. In the latter group clinical improvement was only temporary in the three parathyroidectomized patients. Al hydroxide [Al(OH)3] as a phosphate binder was tapered off in 1981 and substituted by Al-free chelants. In 1987, 66% of patients were given CaCO3 or Mg (OH)2 alone or in association, while 34% still needed Al(OH)3, although at low dosages (less than 2 g/day). The conclusion is that such a protocol is able to prevent and to treat cases of Al intoxication, albeit only partially.

Ribosome maturation in E. coli.

In vivo and in vitro experiments have shown that processing of ribosomal RNA is a late event in ribosome biogenesis. The precursor form of RNA is probably necessary to speed up the assembly of ribomal proteins. Newly formed ribosomal particles which have already entered polyribosomes differ from mature ribosomes not only in their RNA content but also in their susceptibility to unfolding in low Mg concentration and to RNase attack. Final maturation of new ribosomes is probably dependent on their functioning in protein synthesis. Thus only those ribosomes which have proven to be functional may be converted into stable cellular structures.

Maintenance of residual renal function 10 years after the start of hemodialysis: the advantage of tailored schedules?

Maintenance of residual renal clearance is a clinical advantage, protecting against the long-term effects of uremia: although demonstrated in peritoneal dialysis, the strategies in hemodialysis are less clear. This case suggests that dialysis schedules individualized on the basis of renal clearances may help preserve residual function. SB is a 58 year-old male who started dialysis in emergency (creatinine 30.7 mg/dL) in 1993. He had a history of gout, small shrunken kidneys and moderate hypertension. The clinical diagnosis was vasculointerstitial nephropathy. Eighteen months after starting hemodialysis on a conventional thrice weekly schedule, the patient was switched to 2 sessions/week (creatinine clearance increased to 6 ml/min). Thereafter, clearances were checked in alternate months and treatment was tailored to an equivalent renal clearance > or =12 ml/min (1-2 sessions, 2-3.30 hours/week). Ten years after beginning dialysis, he is on a twice weekly schedule (3.30 hours), is normotensive, works full-time and does not want to go on a transplant waiting list.

Hypervitaminosis B12 in maintenance hemodialysis patients receiving massive supplementation of vitamin B12.

We have administered routinely a multivitamin preparation containing a megadose of B12 to 106 hemodialysis patients after dialysis treatments. We found that these patients had very high levels of serum vitamin B12 which returned to original values only after a period of three years after stopping the vitamin. Discontinuing therapy had no effect on hemoglobin, mean erythrocyte corpuscular volume, or motor nerve conduction velocity. It is not known whether maintaining a prolonged high level of vitamin B12 is harmful. However, animal and epidemiologic studies have suggested that both cobalamin and cobalt may be potentially toxic. In view of the absence of demonstrable benefit and the possible risk of toxicity, we believe that the use of such megadose vitamin compounds in dialysis patients should be re-evaluated.

[Complex vascular access]. / Gli accessi vascolari complicati.

Availability of a proper vascular access is a basic condition for a proper extracorporeal replacement in end-stage chronic renal failure. However, biological factors, management and other problems, may variously condition their middle-long term survival. Therefore, personal experience of over 25 years has been critically reviewed in order to obtain useful information. In particular "hard" situations necessitating complex procedures have been examined but, if possible, preserving the peripherical vascular features.

[Use of coil negative pressure in the kidney]. / Utilizzazione della pressione negativa nei reni a rotolo

A sealed compartment recycling system has been created for use with Extracorporeal commercial filters in order to exploit the negative pressure of the dialysate for purposes of ultrafiltration. The results of in vivo and in vitro tests regarding ultrafiltration and dialysates of urea, creatinine, Hipaque I125 and vit. B12 Co57 are reported. The tests highlighted improved dehydration characteristics in the system compared with the traditional coil technique, while the dialysates of small and medium molecules showed no decrease. The system can profitably be used as an alternative to the open compartment coil system and is particularly interesting because it can be combined with monitors which provide exclusively for the use of closed circuit dialysate instruments.

[The quarantined dialysis room for Australia-antigen-positive patients. Theoretical, normative, structural, operational, organizational and management aspects]. / La sala dialisi contumaciale per pazienti antigene Australia positivi. Aspetti teorici, normativi, strutturali, operativi ed organizzativo-gestionali.

The technical features of dialysis facilitate the transmission of hepatitis B virus and the outbreak of possibly serious epidemics among patients and hospital staffs. Statistical, epidemiological and clinical data have been collected on many occasions. They serve to emphasise the problem and the need for quarantined centres. The establishment of such a centre for HBsAg-positive patients is discussed in the light of personal experience in the design and setting up of quarantined rooms. The underlying technical and structural requirements are also examined. Reference is also made to two years' experience in the running of a room of this kind. Stress is laid on the importance of having a room for these patients, not only from the clinical standpoint (lessening of contagion), but also from teh social and exonomic standpoint (reduced costs and length of hospital stay, greater rehabilitation).

[Diabetic nephropathy. Review of the current status and practical considerations concerning etiopathogenetic, clinical and therapeutic aspects]. / La nefropatia diabetica. Rassegna di attualità e considerazioni pratiche di ordine etiopatogenetico, clinico e terapeutico.

Reference is made to personal research and clinical experience, coupled with a select bibliography, in a survey of the most significant current knowledge with regard to diabetic nephropathy, designed for the nonspecialist practitioner and intended to enable him to update his knowledge without plunging into details, and to quickly decide what measures he should adopt in his daily practice. Questions of histopathology and aetiopathogenesis are examined, together with the relations between proteinuria, hypertension, renal function, and diabetic nephropathy. The clinical profile and progress of the disease are illustrated. Lastly, an account is given of the conventional modes of treatment, and of the still partly unsatisfactory results offered by more recent forms of replacement therapy (dialysis and transplantation).

[Hyperparathyroidism in endocrinology and nephrology]. / L'iperparatiroidismo in endocrinologia e in nefrologia.

The literature on primary and secondary hyperparathyroidism is surveyed in the presentation of an account of the present positon. Reference is also made to personal clinical experience, particularly in secondary cases. The physiopathology of parathormone, calcitonin and vitamin D is described; the last, in fact, has proved useful in the treatment of secondary forms. The usual symptomatological tests of performance are explained, together with their diagnostic value. The clinical symptoms and their physiopathological significance are also illustrated. Stress is laid on the importance of conservative management in secondary forms. Good results now enable parathyroidectomy to be avoided or delayed in many cases.