Search

Objective: Tissue transglutaminase was identified as the autoantigen eliciting endomysial antibody. A homemade enzyme-linked immunosorbent assay (ELISA)-based test was recently developed to determine quantitative titers of IgA antitissue transglutaminase antibody. Our objective in this study was to assess the suitability of a newly developed commercial kit for quantitative determination of antibody in patients with untreated celiac disease., Materials: We tested serum samples from 79 untreated celiac patients, 42 healthy blood donors, and 18 patients with nonceliac intestinal disorders evaluated in two different centers. Samples were tested for antitissue transglutaminase, and antiendomysial and antigliadin antibodies in the center where diagnosis was performed. To assess interlaboratory variability of methods, 24 samples randomly selected were blindly tested in both centers. Antitissue transglutaminase antibodies were determined using a commercial kit (INOVA Diagnostics, Inc., San Diego, CA)., Results: Untreated celiac patients had significantly higher titers of antitissue transglutaminase than healthy and disease controls (p < 0.00001). According to the cut-off provided by the manufacturers (20 AU/mL), overall sensitivity was 92% (85% for one center and 100% for the other) and specificity was 98% (100% and 95%, respectively). Antiendomysial antibody was 86% sensitive and 100% specific. Discordance between antitissue transglutaminase and antiendomysial antibodies was detected in 13% of patients. Although two antitissue transglutaminase-negative cases had a positive antiendomysial antibody, the inverse situation was found in eight cases. A blind determination of antitissue transglutaminase on the same samples evidenced a good agreement (kappa statistic: 0.66) between both centers when assessment was qualitative (based on the decision of positive or negative). Although correlation of titers for both determinations was highly significant (r: 0.902, p < 0.00001), a very wide interlaboratory variability (median: 50%) was detected when absolute values were considered., Conclusions: The quantitative determination of antitissue transglutaminase using a commercial kit was highly sensitive and specific for detection of celiac disease. We observed an incomplete overlapping with antiendomysial antibody. The very high variability of values between laboratories still remains to be solved so as to propose the commercial ELISA assay for the screening of celiac disease.

We studied 150 metal workers occupationally exposed to metals and metalworking fluids (MWFs) to determine the prevalence and nature of contact dermatitis. 150 office workers were used as non-exposed control group. Questionnaires were administered to evaluate occupational and non-occupational exposure. All subjects underwent a dermatological examination and patch-testing with standard allergen series and MWFs used in the plant. Twenty-eight metal workers (18.6%) presented minor skin disorders involving the hands (vs. only 2% of the controls), ten (6.6%) had major disorders (similar to the figure for the control group - 5.4%), and 112 (74.8%) had no lesions, as opposed to 92.6% of the control group. Positive patch tests were found in ten metal workers: eight had major skin disorders (six to nickel, cobalt and chromium, one to nickel and cobalt, one to nickel) and the remaining two were asymptomatic (one positive for nickel and chromium, one for nickel). Among the controls there were three cases of positivity, all among asymptomatic subjects. Patch tests with MWFs were negative. The prevalence of dermatoses among the metal workers was significantly higher than that of controls (p<0.01), and all cases of allergy in this group were provoked by metals themselves.

Bovine seminal ribonuclease is a member of the RISBAses (ribonucleases with special biological actions) family. It exerts specific anti-tumor, embryotoxic, aspermatogenic and immunosuppressive activities. The cytotoxic effect of bovine seminal ribonuclease on tumor cells is accompanied by the induction of apoptosis. We provide ultrastructural and flow cytometry evidence of apoptotic death following bovine seminal ribonuclease treatment, in normal cells and phytohemagglutinin-stimulated lymphocytes. Transmission and scanning electron microscopy, which were fully supported by flow cytometry data, showed typical features of apoptosis, such as decreased cell size, chromatin condensation, fragmentation in micronuclei, and the presence of apoptotic bodies.

To assess epidemiological and clinical significance of drug hepatotoxicity in the setting of liver diseases consultation, ten thousand and three hundred forty two prospectively designed clinical records from patient cared for in our Liver Unit in the period 1988-1998 were incorporated into the study; 58 out of 10,342 (prevalence = 5.6%) fulfilled at least the first three of the following causality requirements: 1.--Liver injury associated in time to drug exposition; 2.--Negative evaluation of more common other etiologies; (alcohol, viruses, immunologic, metabolic, etc) 3.--Favourable response to drug withdrawal (ALT < 50% of baseline in 8 to 30 days in acute hepatitis type, and alkaline phosphatase and/or total bilirubin < 50% of baseline up to 6 months, in acute cholestasis) 4.--Inadverted or rarely prescribed positive challenge. Acute hepatitis type of injury were considered when serum ALT rise 8 times or more above normal superior level with alkaline phosphatase (APh) below 3 times; "pure" cholestasis when APh rise 3 times or more above normal with ALT below 8 times; mixed acute injury or cholestatic hepatitis when both ALT and APh were elevated above 8 and 3 times respectively, and indeterminate type when both enzymes were below the referred levels. Chronic injury were considered when six or more month of evolution and compatible liver histology happens. Clinical severity were expressed as mild (absence of major clinical complications, serum bilirubin < 5 mg/dl and prothrombin concentration > 75%), moderate (presence of clinical complications, bilirubin > 5 mg/dl and prothrombin concentration between 50-75%), and severe (major clinical complications with bilirubin > 5 mg/dl and prothrombin concentration < 50%). Female/male ratio was 1.4:1, with age average 39 years (R = 15-77) and major concentration of cases above 40. More than 50% of cases received 2 or more drugs. Jaundice was present in 60.4%, and systemic manifestations of hypersensibility (fever, adenomegalies, rush, mononucleosis like syndrome, eosinophilia) in 29.3%. Acute injury represented 91.4% of the cases: 41.4% acute hepatitis, 15.5% "pure" cholestasis, 24.1% cholestatic hepatitis, and 10.3% indeterminate type. Four patients (4.5% of acute injury cases) were presented as severe acute liver failure, leading to liver transplant in one of them, drug association (INH-rifampicin and carbamazepine-phenobarbital) and inadverted challenge (sulphonamides and pemoline) were associated to clinical severity. Chronic injury were found in five patient (8.6%), four of them associated to chronic hepatitis and the other one to a ductopenic syndrome. Six drugs represented 53.4% of our cases; oral contraceptives (7 cases), INH alone or combined with rifampicin (6 cases), sulfonamides and clorpropamida (5 cases each), carbamazepine and amiodarone (4 cases each). Normalization of liver enzymes after drug suppression took 2 to 8 weeks in acute hepatitis type (X = 4 weeks), 4 to 20 in "pure" cholestasis (X = 12 weeks) and 8 to 24 weeks in cholestatic hepatitis or mixed type (X = 16 weeks). Two cases of chronic hepatitis normalize the histological activity index in 20 and 18 month respectively, one case remains as chronic hepatitis at 10 month and the other one progress to cirrhosis; the ductopenic syndrome normalize histology in 19 months receiving urso-deoxicolic acid, 10 mg/k/day.

Phosphatidylcholine-dependent phospholipase C is an enzyme which hydrolyses phosphatidylcholine giving origin to diacylglicerol and phosphorylcholine. Diacylglicerol has many effect and activates also protein kinase C. Since the presence of protein kinase C in the hepatocyte nuclei and the existence of a phospholipidic fraction in the chromatin have been demonstrated, we investigated if phosphatidylcholine-dependent phospholipase C could be present in the nuclei. The results obtained have shown the presence of this enzyme in the chromatin fraction which differs with respect to that of nuclear membrane in pH and Km. The activity has been also evaluated during liver regeneration. In the chromatin an increase of activity has been shown 12 h and 30 h after hepatectomy, i.e. at the beginning of hepatocyte S-phase. No similar behaviour has been observed in the nuclear membrane. It has been suggested that diacylglicerol, produced by the hydrolysis of chromatin phosphatidylcholine, may have a role in initiating DNA synthesis through the prolonged activation of the nuclear form of protein kinase C., (Copyright 1999 Academic Press.)

Nuclear membrane fluidity is measured in rat liver by use of the fluorescence anisotropy of two probes: diphenylhexatriene and its cationic derivative trimethylammonium-diphenylhexatriene. It has been shown that, in 2-month-old rat liver cells, the bilayer surface is less fluid than the hydrophobic core. The fluidity was higher in 6-day-old rat liver nuclei, in which both the amount of cholesterol and the cholesterol/phospholipid ratio decreased. The influence of the single phospholipids, and in particular of phosphatidylcholine, has been studied by increasing the phosphatidylcholine with a choline base exchange reaction in isolated nuclear membranes. After this reaction, the fluorescence anisotropy of the bilayer surface increased, whereas at the hydrophobic core it decreased. Analysis of fatty acid composition shows an increase of phosphatidylcholine unsaturated fatty acids. The results show that the fluidity of nuclear membranes changes in relation to the lipid content and to the fatty acid composition. The role of nuclear membrane fluidity in cell function is discussed.

In order to have knowledge on the histopathologic characteristics of the HCV infections in our geographical area and its relation with some epidemiologic variables, a series of 54 biopsies of Anti HCV (R) patients was analyzed. The histologic lesions found in this study correspond mostly to patients with relatively early infections, on the contrary to other studies of the some kind. The most frequent histopathologic diagnosis were chronic hepatitis 38/54 (70.4%), steatosis 4/54 (7.4%) and 12/54 (22.2%) with no changes. The presence of lymphoid follicles in the portal tracts was the most frequent histological change in this series (66.7%), followed by the alteration in bile ducts (53.7%); they occurred in a significantly higher proportion in the biopsies which had a diagnosis of chronic hepatitis (p = 0.02) (p = 0.000002). The presence of steatosis and acidophilic bodies in the acinus were found in nearly one third of the biopsies. This findings suggest that the hepatic damage in the anti HCV reactive patients might take place through immunologic mechanisms and cytopathic direct action. It was not found that histologic changes produced by HCV might differ according to epidemiologic variables (post-transfusional, drug abuse i.v. and sporadic).

By color-Doppler ultrasound it's possible to visualize urine flow jet from ureter into the bladder. Aim of the study was to evaluate of ureteral jet in patients with benign prostatic hyperplasia before, during and after with one or two drugs medical therapy. Thirteen patients, aged 51-63 years, were studied; they were not affected by metabolic, hepatic, renal diseases and by prostate inflammation. Eco color Doppler p.w. (Toshiba SSA 270A) with a convex probe of 3.5 MHz was used. A transabdominal ultrasound study was performed, prostate volume measured and ureteral jet visualized before and along treatment (at six months interval) with Finasteride and at the end of treatment. Successively, in four patients, with relapse of prostatic synptomatology, a transabdominal ultrasound study was performed, before and along a treatment with Finasteride, 5 mg/die (Finastid, Neopharmed) and Terazosin hydrochloride, 5 mg/die (Teraprost, Malesci), and at the end of treatment.

The authors report a case of epibronchial esophageal diverticula. The patient underwent a diverticulectomy and subdiverticular myotomy was associated. No significant complications were observed in the post-operative period. In the light of the literature the paper discusses anatomic-clinical, diagnostic and therapeutic aspects of esophageal diverticula.

Spontaneous bacterial peritonitis in patients with chronic hepatitis represents a very controversial matter in terms of frequency, pathology and treatment. This unusual complication mainly due to decreased immunological defences and ascitic fluid opsonic activity is not accepted by all the Authors as a rare event. However, there is agreement as far as management is concerned: cultural examination of the ascitic fluid and the relative antibiogram are the best tools in guiding the approach to an adequate antibiotic therapy.

IL-10 is a cytokine produced by B and T-cells, monocytes and keratinocytes with pleiotropic effects, some of which are directed towards suppressing monocyte activities (anti-inflammatory cytokine). No information at the protein level is available concerning IL-10 in suction blister fluids from psoriatic skin, even if contrasting data have been reported on IL-10 mRNA of psoriatic biopsies and on the cytokine patterns of the T-cell clones, isolated from psoriatic skin. The IL-10 blister fluid concentrations in psoriatic lesions were compared to those found in the non-lesional skin of 14 patients effected with plaque-type psoriasis, and to those found in the skin of healthy controls (9 subjects sharing sex ratio and age with psoriatic patients). No difference in the IL-10 levels was found between non-lesional and control skin. In contrast, lower IL-10 levels were observed in blister fluids obtained from lesional psoriatic skin (p < 0.0005). The possible meanings of these results have been evaluated in the context of the mechanisms activating or maintaining the chronic inflammatory components of psoriasis.

200 divers of either sex, aged 18 to 54 years, entered a double-blind study to compare the efficacy and tolerability of nimesulide 200 mg/day with those of seaprose S 60 mg/day in the treatment of nonbacterial inflammatory disorders of the ear, nose, and throat. At the end of the 1-week treatment period, both drugs were judged to be effective, with improvements and, in most cases, complete remission of all symptoms observed. Nimesulide showed greater clinical efficacy, and both drugs were well tolerated.

Tyrosine aminotransferase activity in rat liver increases during the first 24 hrs after partial hepatectomy with two peaks, one at 10 hrs and another at 18 hrs. This behaviour is due to an increase in TATmRNA synthesis. Expression of serine deydratase is also enhanced during the first 5 hrs after hepatectomy. It is suggested that the enhanced expression of the two genes is due to an increase in hormone incretion particularly glucagon and glucocorticoids.

Tyrosine aminotransferase activity (TAT) and its expression were measured in rats of different ages (3, 12 and 24 months). The enzyme activity showed a circadian rhythm with a peak at midnight due to different levels of transcription during the day. The circadian rhythm was present at all ages studied but showed some differences: an age-related shift of the peak was more evident in the actual levels of transcript. Both specific mRNA and enzyme activity analyses provide better insight into the complex modifications in gene-expression of aging animals.

The aim of this work is to verify, starting from Foulkes' group-analytic perspective, the existence of a link between individual, family and group relational modality on one side, and the many group techniques on the other. This link is well outlined and defined: it is the need to harmonize, to integrate the different parts of the individual, the family, the group. The satisfaction of this need leads to well-being; its dissatisfaction leads to disorders and to the consequent relational modality. In particular, the lack or the reduction of the integrative and associative capacity leads to psychosis, while the disharmony of the different psychical functions leads to neurosis, and the body disharmony leads to psychosomatic disorders. Following Foulkes' group-analytic perspective, the author works out a new definition of man. Man is the final function of all the body functions.

We used RNAase protection and restriction fragment length polymorphism assays to detect activating mutations of c-src in a spectrum of human tumours. No mutations were detected at codons 98, 381, 444, and 530. We conclude that mutational activation is not the mechanism of enhancement of pp60c-src-specific kinase activity found in a number of human cancer types.

The activity of the enzyme tyrosine aminotransferase and the synthesis of its specific mRNA were evaluated at different hours of the day in the liver of 3-, 12- and 24-month old BN rats. The enzyme activity has a circadian rhythm with a peak at midnight in 3- and 12-month old, which shifts to 03.00 hrs in 24-month old animals, in agreement with previous results. The expression of TATmRNA also changes during the day indicating circadian fluctuations which change with age. In 3-month old rats the TATmRNA peak is at 19.00 hrs, preceding that of the enzyme activity. In 12-month old rats the TATmRNA synthesis reaches a maximum at midnight and in 24-month old rats at 03.00 hrs. The results show that the circadian rhythm of tyrosine aminotransferase activity is due to a different gene expression throughout the day, which is influenced by age.

We describe a simple procedure to measure precisely the tip diameter of glass micropipets. This procedure can improve the precision of microinjection and some electrophysiological measurements due to their extreme sensitivity to the pipet's exact geometric dimensions. The technique is nondestructive and readily carried out as a brief, extra step in these applications. Digital measurements of the threshold pressures for gas bubbling from (gas filled) pipets submerged in methanol were used to calculate the corresponding inner tip diameters using the LaPlace equation. Direct measurements of the inner tip diameters (from 0.2-5.0 mu) using scanning electron microscopy confirmed the validity of this indirect, simple procedure.

To verify the level of participation of children and teachers in an educational program, the reliability of the program itself and to determine whether children can modify family's behavioral patterns, a feasibility study was undertaken among the pupils of a primary school. Seventy-two children, belonging to different socio-economic classes, were studied. Children were given a questionnaire before and after the course, which was held by previously trained teachers. Very good participation of children and teachers in the program was observed. On the contrary very low involvement of the parents was achieved. This experience emphasizes the difficulties arising when actions to modify adult's life styles are attempted. On the contrary, a standardized education program, lasting for a whole school year, could be a feasible, useful and not too costly program not only for the primary prevention of coronary artery disease, but for any program of extensive health care.

Axonal transport of phospholipids in normal and regenerating sciatic nerve of the rat was studied. At various intervals after axotomy of the right sciatic nerve in the midthigh region and subsequent perineurial sutures of the transected fascicles, a mixture of 60 mu Ci [Me-14C]choline and 15 muCi [2-3H]glycerol in the region of the spinal motor neurons of the L5 and L6 segments was injected bilaterally. The amount of radioactive lipid (and in certain cases its distribution in various lipid classes) along the nerve was determined as a function of time. Three days after fascicular suture and 6 h after spinal cord injection of precursors, there was an accumulation of labeled phospholipids and sphingolipids in the transected sciatic nerve in the region immediately proximal to the site of suture. Nine days after, there was a marked increase in the accumulation of radioactivity in the distal segments of the injured nerve, which increased up to 14 days after cutting and disappeared as regeneration proceeded (21-45 days). In all segments of both normal and regenerating nerve fibers, as well as in L5 and L6 spinal cord segments, only phosphatidylcholine and sphingomyelin were labeled with [14C]choline. These results suggest that the regeneration process in a distal segment of a peripheral neuron, following cutting and fascicular repairing by surgical sutures, is sustained in the first 3 weeks by changes in the amount of phospholipids rapidly transported along the axon towards the site of nerve fiber outgrowth.

The content of alkenyl-acyl, alkyl-acyl and diacyl types of the three major myelin glycerophospholipids such as PtdCho, PtdEtn and PtdSer was determined in myelin fractions prepared from sciatic nerve segments of rats at 12, 25 and 45 days after birth, and of adult rats (6-month-old) 90 days after crush injury. The biosynthesis and metabolic heterogeneity of lipid classes and types were also studied by incubation with [1-14C] acetate of nerve segments of young rats at different ages as well as crushed and sham-operated control nerve segments of adult rats. The analysis of composition and positional distribution in major individual molecular species extracted from light myelin and myelin-related fraction suggest that the metabolism of alkenyl-acyl-glycerophosphorylethanolamines and unsaturated species of PtdCho and PtdSer may not be regulated in the same manner during peripheral nerve myelination of developing rat and remyelination of regenerating nerve in the adult animal. The 14C-radioactivity incorporation into lipid classes and alkyl and acyl moieties of the three major phospholipids of sciatic nerve segments during the developmental period investigated revealed that Schwann cells were capable of synthesizing acyl-linked fatty acids in both myelin fractions at a decreasing rate and with different patterns during development. In regenerating sciatic nerve of adult animals the labeling of myelin lipid classes and types of remyelinating nerve segment distal to the crush site was markedly higher than that of sham-operated normal one; however, the magnitude and the pattern of the specific radioactivity never approached those observed during active myelination of the nerve in young animals.(ABSTRACT TRUNCATED AT 250 WORDS)

Personal experience of liver surgery for benign and malignant pathology is reported. The type of operation and results obtained in relation to mortality and possible intra- and postoperative complications are considered. The extraparenchymal and transparenchymal techniques are both considered valid but in approaching the vascular structures the former is preferred because it permits preliminary vascular monitoring. The results obtained are similar to those of other series reported in the literature.

The concept of a subtractive therapeutic intervention in several diseases (shock, acute liver, failure, poisoning) to remove toxic middle molecules (MM) has led to the devising of a particular ultrafiltration system (anadiafiltration) to clear weakly bound MM. Clinical hemoanadiafiltration (HADF) with a polyacrylonitrile filter and in vitro experiments with a series of marker substances, have shown HADF efficacy in clearing MM, irrespective of their molecular weight, with protein binding K between 50 and 150.

The contribution of the axonal transport to the biosynthesis of myelin phospholipids was investigated in the rabbit optic pathway. A double labeling technique was used. The same animals were injected with one isotope intravitreally and the other intraventricularly. This procedure allows double labeling of the optic nerves, optic tracts, lateral geniculate bodies (LGB), and superior colliculus (SC). The precursors simultaneously injected were: [1-14C]palmitate (15 microCi intravitreally in both eyes or 50 microCi intraventricularly) and [2-3H]glycerol (50 microCi intravitreally in both eyes of 100 microCi intraventricularly). Twenty four hours and 10 days after the injections, myelin was purified from pooled optic nerves and optic tracts as well as from pooled LGBs or SCs. The phospholipids were extracted and then separated by thin-layer chromatography; the specific radioactivity of the various classes of phospholipids was determined. Using both administration routes of C- or 3H-precursors, the distribution of label and specific radioactivity of myelin phospholipids in the retina and in all other optic structures were very similar. Phosphatidylcholine, phosphatidylethanolamine and phosphatidylserine + phosphoinositol were preferentially labeled with both precursors. These results suggest that, in the rabbit optic pathway the phospholipids synthesized in the retinal ganglion cells and transported along the axons, could undergo transaxonal transfer into myelin.

The protein products (p21) of the ras cellular proto-oncogenes are thought to transduce membrane signals necessary for the induction of cell division. However, there is uncertainty as to the precise role of ras p21 in mediating ligand-membrane receptor signals leading to cell differentiation. Treatment of rat phaeochromocytoma cells (PC12) with nerve growth factor (NGF) results in the induction of a number of phenotypic characteristics of sympathetic neurones, including cessation of cell division and outgrowth of neuronal processes (neurites). Here we report that microinjection of antibody to ras p21 into PC12 cells inhibited neurite formation and resulted in temporary regression of partially extended neurites, an effect which was observed up to 36 h after initiation of NGF treatment. Neurite formation induced by cyclic AMP was unaffected by injection of anti-p21 antibody. These results indicate that p21 is involved in the initiation phase of NGF-induced neurite formation in PC12 cells and has a role in hormone-mediated cellular responses distinct from cell proliferation.

Changes of lipid, free fatty acid, protein, DNA, and RNA content in proximal and distal segments of regenerating sciatic nerve, from 14 to 120 days after crush, were determined. During the early stage of Wallerian degeneration, a marked decrease of phospholipid, cerebroside and sulfatide content and, in contrast, a marked increase of protein, DNA, RNA, and free fatty acid content, in the distal segment of crushed nerve compared to control, was observed. A gradual increase of phospholipid, cerebroside, and sulfatide levels, approaching normal values, and a gradual slope in the increase of protein, DNA, RNA, and free fatty acid levels over the ensuing time periods of regeneration was seen. Total cholesterol content was relatively constant during regeneration, slightly increasing at day 120. The activity of 2', 3'-cyclic nucleotide 3'-phosphodiesterase (CNPase) of myelin fraction purified from distal segment of regenerating sciatic nerve showed a significant increase in the 30-120 day regenerating period. A marked increase of the incorporation of [2-3H]glycerol and of [Me-14C]choline into myelin lipids of distal segment of regenerating nerve, was found. Labeling of myelin lipids with [3H]oleic acid (injected intravenously seven days before crush) support the evidence that a similar pattern of degeneration exists between two different types of trauma, i.e. nerve crush or cut. The findings suggest that, in the distal segment of crushed nerve, the lipid content as well as the myelin lipid synthesis increase as the regeneration period proceeds.

The labeling of RNA in young and adult rat brain has been studied by measuring in vitro (tissue slices incubation) the incorporation of labeled uridine into RNA of total tissue and of the various subcellular fractions purified from cerebral hemispheres of 1- and 10-month-old rats. Gel electrophoretic analysis of the newly synthesized nuclear and microsomal RNA was also accomplished. An active metabolism of RNA in adult animals was found; moreover, distinct differences in ribosomal RNA processing in cerebral hemispheres of 1- and 10-month-old rats, with a more rapid processing in the brain of adult animals, were obtained.

The uptake of different labeled precursors, their incorporation into lipids, and transport along the rabbit optic pathway [ipsilateral retina and optic nerve (ON), and contralateral optic tract (OT), lateral geniculate body (LGB), and superior colliculus (SC)] were investigated. Albino rabbits were used. The following radioactive precursors ,either combined or separately, dissolved in 50 microliter of saline containing 15% BSA, were injected into vitreous body: [2-3H]glycerol (50 microCi), [1-14C]palmitate (15 microCi), and [1-14C]linoleate (7.5 microCi). Animals were killed at different time intervals from 1 hr up to 24 days. The radioactivity of total lipids and of different phospholipid classes from total tissue was measured. One hour after administration of precursors, the radioactivity into the retina was high and the incorporation of [3H]glycerol and [14C]palmitate increased until 12 hr and 24 hr, respectively. The incorporation of [14C]linoleate reached a maximum on the second day. The phospholipids of LGB and SC were intensively labeled after 4-8 hr, and their radioactivity increased up to the 10th day after injection, independent of the precursor employed. The results obtained indicate that the labeled hydrophilic and hydrophobic precursors used were actively incorporated into the retina, The phospholipids were later transported at a rapid rate along the optic pathway.

We examined the effects of age on RNA and lipid formation by whole retina and optic nerve in vitro. Male Wistar rats, aged 4, 12, and 24 mo, were used. From the results obtained the following conclusions may be drawn: 1. In assaying the lipid biosynthesis during aging, a striking difference between the retina and optic nerve clearly emerged; 2. In isolated retina, [3H]uridine incorporation into RNA was relatively constant at the three ages, whereas both [14C]palmitate and [3H]choline incorporation into lipids showed a substantial increase in rats at 24 mo of age compared with those at 4 mo; 3. In contrast, in the optic nerve of the oldest rats, compared with the youngest, a significant decrease of [14C]acetate and [14C]palmitate incorporation into acylglycerols, cerebrosides, and phospholipids was found. Each fatty acid precursor label was incorporated to a proportion that reflected the typical acyl group composition of individual lipids; 4. Following labeling of the optic nerve with [3H]choline, the specific radioactivity of choline-containing phospholipids was drastically decreased with increasing rat age; and 5. The incorporation of [2-3H]glycerol into optic nerve diacylglycerols, PtdEtn, and PtdIns declined with age, whereas no significant change took place in the incorporation into PtdCho. The results strongly support the concept that RNA metabolism of rat retina (most likely photoreceptor cell layer) is not altered during aging; on the contrary, phospholipid synthesis is stimulated in comparison with that of the optic nerve, for which a serious impairment was concomitantly observed. The physiological significance of these responses, and the mechanism by which retinal tissue is spared from the general age derangement of the nervous system, remain to be defined.

The effect of CDP-choline on the in vivo incorporation of labeled precursors into DNA, RNA, and proteins in cerebral hemispheres, cerebellum, and brainstem of guinea pigs after hypoxic treatment was studied. The labeling of macromolecules extracted from the various subcellular fractions of these brain regions was also determined. Hypoxic treatment affected macromolecular labeling to a different extent in the three brain regions examined. CDP-choline treatment was not able to reverse the effect of hypoxia on DNA labeling, but it was able to remove the effect of hypoxia on RNA and protein labeling. The action of CDP-choline was particularly evident on the labeling of RNA in nuclei and mitochondria of the cerebellum and on the labeling of proteins in microsomes of the three brain regions examined.

In the present work, the hypothesis that the increased rapid intracellular transport of newly-synthesized material along the axons of a regenerating system is sustained by an alteration of the transport of proteolipid complexes through subcellular compartments of a neuronal cell body was tested by a biochemical methodology. The motoneurons of spinal cord ventral horn, 4 wk after unilateral lesion (crush) of cervico-thoracic nerves of the rabbit at the level of brachial plexus, were chosen as the model system of regeneration. A time-staggered procedure of in vivo and in vitro double labeling with metabolic precursors, such as [3H]-choline, [14C]-choline, [3H]-fucose, and [14C]-fucose, was used. Subcellular fractions (RER, SER, Golgi apparatus, and plasma membranes) of ventral horn tissue, taken from spinal cord hemisections (regenerating and contralateral side), were further isolated. Twenty-eight days after axotomy, we did not observe any change of intracellular transport kinetics (14C/3H ratio) of newly-synthesized choline-phospholipids and glycoproteins in regenerating motoneurons compared to controls. However, associated with regenerating phenomenon in Golgi apparatus, we observed an increase of labeled choline-phospholipid and glycoprotein material that could contribute to the increased fast axonal transport and delivery of membrane proteolipid complexes to plasma membrane and axonal compartments. The increase of glycoprotein labeling was more pronounced in the SER portion (vesicles and elements of smooth membranes). This result is in favor of the hypothesis that membrane-bound proteins are transported from the Golgi to the axon through the perikaryal SER.

The incorporation of [methyl-3H]thymidine into DNA, of [5-3H]uridine into RNA, and of [1-14C]leucine into proteins of cerebral hemispheres, cerebellum, and brainstem of guinea pigs after 80 hr of hypoxic treatment was measured. Both in vivo (intraventricular administration of labeled precursors) and in vitro (tissue slices incubation) experiments were performed. The labeling of macromolecules extracted from the various subcellular fractions of the above-mentioned brain regions was also determined. After hypoxic treatment the incorporation of the labeled precursors into DNA, RNA, and proteins was impaired to a different extent in the three brain regions and in the various subcellular fractions examined; DNA and RNA labeling in cerebellar mitochondria and protein labeling in microsomes of the three brain regions examined were particularly affected.

[14C]palmitate and [3H]choline were injected intravitreally and, at the same time, intraventricularly in Wistar male rats at 4, 10, and 24 mo of age. The precursor incorporation into lipids of the retina, optic nerve tract, superior colliculus, and lateral geniculate body was followed for 2 h. The specific radioactivity of precursors pool (choline, phosphorylcholine, and free fatty acids) showed a marked decrease in optic nerve tract and lateral geniculate body of aged rats, whereas in retinal tissue and superior colliculus no changes were observed as a function of age. In rats of the three age groups, whole retina and superior colliculus showed neither changes of choline incorporation into phosphatidylcholine and sphingomyelin nor alteration of palmitate incorporation into diacylglycerols, triacylglycerols, and major phospholipid classes as a function of age. In sharp contrast, the optic nerve tract and, to a lesser extent, the lateral geniculate body exhibited a significant age-related decline of either the incorporation of both precursors into all lipid classes or the specific radioactivities of endogenous precursor pools. We concluded that the visual pathway structures are metabolically affected in a different manner by aging. Particularly, the ability of the retina and superior colliculus to metabolize lipids appeared to be age invariant. The marked decline of lipid biosynthesis with age, for some visual structures, is consistent with the trend generally observed in metabolic turnover and function of other CNS regions.

Myelin protein synthesis, as well as ultrastructural and morphometric changes in regenerating peripheral nerve, was studied. Sciatic nerves of rats were crushed unilaterally; sham-operated nerves of the contralateral side served as controls. For the in vivo experiments, rats were killed at selected periods after the nerves were crushed (30, 60, 90, and 120 days); seven days prior to killing, the animals were injected intravenously with L-[4,5-3H]leucine. For the in vitro experiments, proximal and distal segments of sciatic nerve and equivalent sham-operated nerves were labeled with 3H-amino acid mixture 90 days after axotomy. Purified myelin was isolated from nerve segments; specific radioactivity and gel electrophoretic patterns of proteins were analyzed. Cross-sectional electron microscope (EM) preparations of proximal, distal, and contralateral segments of nerves also were examined. Results showed that the incorporation of labeled amino acids into total myelin proteins was enhanced significantly in the distal segment of sciatic nerves at all of the periods of regeneration studied. The yield of myelin protein per mm distal nerve segment increased as regeneration proceeded. The remyelination of fibers early after nerve crush was weak, whereas it gradually attained the normal range 90-120 days after axotomy. Morphometric analysis of myelin sheath thickness of regenerating axons was consistent with the data obtained for myelin protein synthesis.