Human lymphoblastoid cell lines secreting antibodies with restricted HLA specificity

Peripheral B lymphocytes obtained from three healthy individuals who had been immunized against peripheral blood lymphocytes from appropriate HLA-incompatible donors were transformed by the use of Epstein-Barr virus. The transformed blastoid B cells were repeatedly subcultured by means of cluster picking, and the HLA antibody-producing cultures were identified by testing the culture supernatants by means of the cytotoxicity assay, using the corresponding donor cells. Thus far, four cell lines that secrete cytotoxic HLA antibodies (MP1, 3, 4, and 5) have been established. Specific immunoabsorption experiments revealed that the antibody activity is carried by lambda-type IgM for MP1, by kappa-type IgM for MP3 and MP5, and by both for MP4. Specificity analysis of a panel of HLA-pretyped cells indicated that MP1 detects DQw2, whereas MP5 recognizes B7. The specificity of MP3 was similar to a DQ specificity termed DC5 (probably equivalent to TA10) but not the same. In the case of MP4, both of the lambda-type and kappa-type antibodies appeared to be directed toward new HLA class 11 determinants.Abbreviations used in this paper HLA human major histocompatibility - EBV Epstein-Barr virus - B-LCL Blymphoblastoid cell line - NA not absorbed - PBS phosphate-buffered saline - SPA Sepharose protein A - NRS normal rabbit serum     

DNA typing of HLA-DR β chain genes can discriminate between undetected alleles and real homozygotes

The polymorphism of HLA-DR antigens has been studied by Southern blot hybridization under conditions specific for the detection of the DR chain genes. Haplotype-specific patterns were defined with DNA from DR1, 2, 3, 4, 7, w8, w11, w12, and W13 homozygous typing cells, with restriction enzymes Eco RI, Bgl I, and Pvu II. Certain serological specificities, such as DR2, DR3, and DR7, can be encoded by distinct allelic forms of DR chain genes. The procedure of DNA typing was applied to family analysis of individuals expressing only a single DR specificity upon serological typing. Three cases are described here: (1) in family GR, phenotypic DR 7 homozygotes correspond to genomic heterozygotes, and a novel DR7 allele is described: (2) in family RU, the genes corresponding to a serologically undetected (blank) DR allele were identified by restriction fragment length polymorphism (RFLP); this novel DR haplotype has an RFLP pattern similar to those of the DRw52 family, even though this specificity was not expressed on the DR-blank lymphocytes; (3) in family RG, there is no blank allele, but a homozygote RFLP situation at the DR subregion.     

Influence of Hydrocortisone on Chick Embryo Retina Development

Treatment of chick embryos in ovo with hydrocortisone-21-phosphate (a single dose of 150 micrograms) caused a marked reduction of retinal thymidine kinase activity 24 h later. The inhibitory effect was highest (65-70%) in 8-10-day-old embryos and declined with age, disappearing after day 15. It was accompanied by a reduction in thickness of the retinal layers. Adrenocorticotropic hormone (ACTH) treatment (10 micrograms daily for 2 days) also produced an age-dependent inhibitory effect on retinal thymidine kinase, whereas treatment with a single dose of 200 micrograms of metopirone, a compound that prevents the 11 beta-hydroxylation of steroid molecules in the adrenal glands, impeded the decrease in thymidine kinase activity that normally occurs in chick embryo retina after day 9 of development. In addition, metopirone prevented the inhibition exerted by ACTH on thymidine kinase activity but had no effect on the action of hydrocortisone.     

Stereological and functional investigations on isolated adrenocortical cells: Zona fasciculata/reticularis cells of chronically ACTH-treated rats

Summary The morphology and function of isolated inner (zona fasciculata/reticularis) adrenocortical cells of rats pretreated with ACTH for 3, 6, 9 or 12 days were investigated. ACTH treatment induced a notable time-dependent enhancement in the steroidogenic capacity (corticosterone production) and growth of inner cells. The volumes of cells, mitochondrial compartment, membrane space [the cellular space occupied by smooth endoplasmic reticulum (SER) membranes] and lipid-droplet compartment, as well as the surface area of mitochondrial cristae and SER tubules, were increased in relation to the duration of ACTH pretreatment, and showed a highly significant positive linear correlation with both basal and stimulated corticosterone production. The acute exposure of isolated cells to ACTH provoked a striking lipid-droplet depletion, the extent of which was linearly and positively correlated with stimulated corticosterone secretion. The hypertrophy of the mitochondrial compartment and SER are interpreted as the morphological counterpart of the enhanced steroidogenic capacity of inner adrenocortical cells, inasmuch as the enzymes of steroid synthesis are located in these two organelles, and it is well known that chronic ACTH exposure stimulates the de novo synthesis of many of them in vivo. The rise in the number of lipid droplets, in which cholesterol is stored, is interpreted as being due to the fact that, under chronic ACTH treatment, the processes leading to cholesterol accumulation in adrenocortical cells (exogenous uptake and endogenous synthesis) exceed those of its utilization in basal steroid secretion. Cholesterol accumulated in lipid droplets as a reserve material may be rapidly utilized after acute ACTH exposure to meet the needs of the enhanced steroidogenic capacity of adrenocortical cells.     

Ten-Year-Replicated Circadian Profiles for 36 Physiological, Serological and Urinary Variables in Healthy Men

At 3-hr intervals over a 24-hr span, 36 systemic, serologic and urinary variables were examined in 7 men in their mid 20's in the Spring of 1969, and again in the same 7 men in the Spring of 1979 under a similar chronobiologic protocol, using the same chemical and numerical analytical procedures. The variables examined for rhythms by cosinor were: vital signs--blood pressure (systolic, diastolic, pulse pressure and mean arterial pressure), heart rate, intraocular pressure (left and right), oral temperature; serum components--albumin, albumin/globulin ratio, total bilirubin, calcium, carbon dioxide, chlorides, bilirubin, cholesterol, globulin, glucose, potassium, sodium, sodium/potassium ratio, transaminase, triglycerides, total protein, urea nitrogen; and urine components--calcium, calcium/magnesium ratio, creatinine, magnesium, pH, potassium, sodium, sodium/potassium ratio, urea clearance, urea nitrogen, volume and zinc. Although all subjects appeared clinically healthy in 1969 and in 1979, certain inter-study differences were observed in a number of rhythm parameters of different variables. Statistically significant increases in mesor for the group as a whole were observed for serum Ca, cholesterol, Cl, CO2, K, Na, and while statistically significant mesor decreases for a group as a whole were noted in serum glucose and transaminase. Statistically significant increases in amplitude for the group as a whole were observed in serum chloride and urinary Na/K ratio, while statistically significant decreases were observed in amplitude for blood pressure, heart rate, serum albumin, A/G ratio, globulin, glucose, protein, sodium and transaminase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of prolonged sodium restriction on the morphology and function of rat adrenocortical autotransplants

Summary Regenerated adrenocortical nodules were obtained by implanting fragments of the capsular tissue of excised adrenal glands into the musculus gracilis of rats (Belloni et al. 1990). Five months after the operation, operated rats showed a normal basal blood level of corticosterone, but a very low concentration of circulating aldosterone associated with a slightly increased plasma renin activity (PRA). Regenerated nodules were well encapsulated and some septa extended into the parenchyma from the connective-tissue capsule. The majority of parenchymal cells were similar to those of the zonae fasciculata and reticularis of the normal adrenal gland, while zona glomerulosa-like cells were exclusively located around septa (juxta-septal zone; JZ). In vitro studies demonstrated that nodules were functioning as far as glucocorticoid production was concerned, while mineralocorticoid yield was very low. Prolonged sodium restriction significantly increased PRA and plasma aldosterone concentration, and provoked a marked hypertrophy of JZ, which was due to increases in both the number and average volume of JZ cells. Accordingly, the in vitro basal production of aldosterone and other 18-hydroxylated steroids was notably enhanced. The plasma level of corticosterone, as well as zona fasciculata/reticularis-like cells and in vitro production of glucocorticoids by regenerated nodules were not affected. These findings, indicating that autotransplanted adrenocortical nodules respond to a prolonged sodium restriction similar to the normal adrenal glands, suggest that the relative deficit in mineralocorticoid production is not due to an intrinsic defect of the zona glomerulosa-like JZ, but is probably caused by the impairment of its adequate stimulation under basal conditions. The hypothesis is advanced that the lack of splanchnic nerve supply and chromaffin medullary tissue in regenerated nodules may be the cause of such an impairment.     

Serological discrimination of DR4 haplotypes by radioimmunoassay

Three new alloantigenic specificities of human major histocompatibility complex class 11 molecules have been defined by testing the reactivity of alloantisera at the molecular level. Two of these specificities identify different DR4 haplotypes. The Fe75 specificity is associated with the DR4/DW10 haplotype and the CBC/MRG6 specificity with the DR4/DKT2 haplotype. Both are supertypic specificities and are associated with other DR specificities as well. Both specificities are carried by class 11 molecules belonging to the first DR subset. Together with previously described determinants, these specificities contribute to serological discrimination of the different DR4 haplotypes.     

Neuroendocrine modulation of haloperidol-induced catalepsy

Evidence has been accumulated implicating sex hormones as possible modulators of extrapyramidal motor function. In the present study we have investigated the effects of estrogens, progesterone, testosterone, prolactin and calcitonin on behavioral parameters related to nigro-striatal dopaminergic system, such as haloperidol-induced catalepsy in male rats. It was found that 7-days estradiol benzoate treatment (5 micrograms/rat/day) significantly increases haloperidol-induced catalepsy, suggesting a possible antidopaminergic activity of estrogens. On the other hand, prolactin facilitates nigro-striatal dopaminergic transmission. Interestingly, 7 day treatment with medroxy-acetate progesterone (MAP, 5 mg/Kg, i.p.) brings about a trend to a decrease in haloperidol-induced catalepsy, while no significantly effect was observed following acute MAP administration at the same dose. So, it is tempting to speculate that chronic progestinic treatment may result in an increase in dopaminergic tonus. Testosterone, acutely administered (5mg/kg.s.c.) induces changes similar to those observed following progesterone administration. Finally, also calcitonin is able to influence haloperidol-induced catalepsy by markedly increasing it.