PJA-BP expression and TCR delta deletion during human T cell differentiation

Recombination of deltaRec to psiJalpha will delete the TCR delta gene, which is thought to play an important role in the bifurcation of the TCR alphabeta versus TCR gammadelta differentiation lineages. We recently detected a DNA-binding protein in human thymocytes, the so- called PJA-BP, which recognizes the psiJalpha gene segment and might be one of the factors involved in the regulation of preferential deltaRec- psiJalpha rearrangements. We now investigate PJA-BP expression and its correlation with TCR delta gene deletion in thymocytes. Our electrophoretic mobility shift assay experiments showed that the PJA-BP is evolutionary conserved in human, murine and simian thymocytes. Using a large series of human hematopoietic malignancies (n = 30), we conclude that PJA-BP expression is thymocyte specific and seems to be restricted to thymocytes committed to the TCR alphabeta lineage. Analysis of seven well-defined human thymocyte subpopulations showed that preferential deltaRec-psiJalpha rearrangements as well as PJA-BP expression can be detected from the immature CD34-/CD1+/CD3- /CD4+/CD8alpha+beta- thymocyte differentiation stage onwards. These experiments indicate that expression of PJA-BP in human thymocytes starts simultaneously with preferential deltaRec-psiJalpha rearrangements, which supports our hypothesis that PJA-BP is one of the factors involved in the preferential recombination of deltaRec to psiJalpha.      

Molecular genetic analysis of autosomal dominant cerebellar ataxia with retinal degeneration (ADCA type II) caused by CAG triplet repeat expansion

Autosomal dominant cerebellar ataxia with retinal degeneration (ADCAII) was previously mapped by linkage analysis studies to chromosome 3p12- p21.1 (SCA7). Positional cloning efforts have recently identified a novel gene, SCA7 , containing a translated CAG repeat, expanded in SCA7 patients. We cloned the SCA7 gene from a yeast artificial chromosome (YAC) clone contig spanning the SCA7 candidate region. Using a combination of genomic sequencing and cosmid-based exon trapping, two expressed sequence tags were identified. Sequencing of the corresponding cDNA clones and RT-PCR analysis identified the full- length SCA7 cDNA. Together, our sequence data defined the intron/exon boundaries of the first two coding exons of the SCA7 gene, with the first exon containing the expanded CAG repeat. Further, sequence comparison with the published SCA7 cDNA identified one additional putative exon in the 5'-UTR region of the SCA7 gene. The SCA7 gene was mapped on the YAC contig in the 2.5 cM interval between D3S1600 and D3S1287. In one extended Belgian SCA7 pedigree the expanded alleles ranged from 38 to at least 55 repeats with allele lengths being inversely correlated with onset age of ADCAII symptoms. The SCA7 repeats increased in length in successive generations. Normal alleles had from four to 18 repeats, with 10 repeats being the most common allele.      

Effects of intracerebral implants of steroid hormones on scent marking in the ovariectomized female gerbil (Meriones unguiculatus)

The effects of intracerebral implants of steroid hormones on scent marking in the female gerbil (Meriones unguiculatus) were studied in two experiments. In Experiment 1 various steroids were implanted alone or in combination into the preoptic and anterior hypothalamic area of ovariectomized females. Unilateral implants of testosterone + estrogen, estrogen, estrogen + progesterone, testosterone and testosterone + progesterone stimulated a significant level of marking when compared to controls. Experiment 2 utilized bilateral implants of estrogen dissolved in paraffin in order to explore the distribution hormone sensitive areas in the brain which might be important in the regulation of scent marking in the female gerbil. Pellets of estrogen-paraffin were implanted stereotaxically into either the anterior hypothalamus, preoptic area, septum, hippocampus, thalamus, amygdala or anterior olfactory nucleus of ovariectomized females. Total dosage of hormone implanted was 8.2–8.4 μg. A significant level of marking resulted in animals receiving implants into the anterior hypothalamus, preoptic area and septum when compared to controls. Marking appeared at about the same rate in each of these groups; however, the level of marking attained differed. By the last trial, anterior hypothalamic implanted animals were marking significantly more often than animals in either the preoptic or septum groups. Although there was no evidence of ieakage from the brain, the data suggested that some rapid diffusion of hormone, largely restricted to the brain, was taking place or that the three areas were differentially responsive to the hormone. The data do indicate that some localization of function does exist with respect to regulation of scent marking in the female.     

Blood supply of the olfactory nerve

Summary The authors report the results of a series of dissections and anatomic sections of the fronto-basal region of the brain and of the anterior cranial fossa in human cadavers. The constant presence of an arachnoidal cistern above the olfactory nerve was verified. The arachnoid separates from the pial membrane and forms a bridge with the ventral part of the olfactory bulb and tract, from the lateral edge of the olfactory sulcus to the medial edge of the gyrus rectus. The cistern is wide in its anterior portion, between the gyrus rectus and the olfactory bulb, and is reduced to a virtual slit in its posterior portion where the tract is lodged in the olfactory sulcus. The olfactory nerve can be separated without damaging fronto-basal arachnoidial adhesions over several centimeters. Dissection of this region after intravascular injection of colored media shows the constant presence of an artery destined to the olfactory bulb and tract. It originates either from the lateral surface of the anterior cerebral a. (segment A2), or from the medial fronto-basal a., and consistently provides terminal branches in front of the olfactory trigone in the medial olfactory sulcus. At their ventral extremity, the olfactory structures are therefore vascularised independently for several centimeters, from the lower face of the frontal lobe. The independent vascularisation of the olfactory nerve, the tenuous and easily detachable adhesions, and the actual presence of a true arachnoidal cistern all contribute to enabling surgical techniques which conserve olfactory function during anterior approaches.

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Vascularisation du nerf olfactif. Rapports méningés et applications chirurgicales

Résumé Les auteurs rapportent les résultats d'une série de dissections et de coupes de la région fronto-basale de l'encéphale et de la fosse crânienne antérieure sur sujets cadavériques. La présence constante d'une citerne arachnoïdienne au dessus du n. olfactif a été vérifiée. L'arachnoïde se sépare du feuillet pial et passe en pont à la partie ventrale du bulbe et du tractus olfactifs, du bord latéral du sillon olfactif au bord médial du gyrus rectus. La citerne est large dans sa portion antérieure, entre le gyrus rectus et le bulbe olfactif, se réduit à une fente virtuelle postérieure lorsque le tractus se loge dans le sillon olfactif. Le n. olfactif peut être séparé sans dommage des adhérences arachnoïdiennes fronto-basales sur quelques centimètres. La dissection de cette région, après injection intravasculaire de masses colorées montre, de façon originale, la présence constante d'une artère destinée au tractus et au bulbe olfactifs. Elle naît soit de la face latérale de l'a. cérébrale antérieure (segment A2), soit de l'a. fronto-basale médiale, pour donner ses branches terminales toujours en avant du trigone olfactif dans le sillon orbitaire médial. Sur quelques centimètres à leur extrémité ventrale, les structures olfactives ont donc une vascularisation indépendante de la face inférieure du lobe frontal. L'indépendance vasculaire du n. olfactif, des adhérences ténues, facilement détachables, et la réalité vérifiée d'une véritable citerne arachnoïdienne permettent d'imaginer des techniques conservatrices de la fonction olfactive utilisées dans plusieurs indications de la chirurgie de la fosse crânienne antérieure.

     

The infrapyloric artery and cephalic pancreatoduodenectomy with pylorus preservation: preliminary study

Summary Cephalic pancreatoduodenectomy (CPD) with pylorus preservation has been suggested to improve the functional and nutritional result of surgery. At operation, the first two centimeters of the duodenum are preserved, the vascular arch of the lesser gastric curvature is saved and the right gastroepiploic artery is resected at its origin. The aim of this study on 15 fresh cadavers was to determine the origin of the vascularization of the remaining duodenum and also the possibilities of preserving an optimal vascularization after CPD and pylorus preservation. All of the arteries supplying the remaining duodenum and arising either from the right gastric artery or the right gastroepiploic artery were identified. The distances between the origin of the infrapyloric artery and the termination of the gastroduodenal artery on the cranial and ventral pancreaticoduodenal artery and the left gastroepiploic artery were measured. At CPD with pylorus preservation, the study demonstrated that: 1) the cranial side of the remaining duodenum remains vascularized in 80% of the cases by one or two supraduodenal branches coming from the right gastric artery; 2) ligation of the right gastroepiploic artery eliminates all vascular supply to the caudal side of the remaining duodenum in almost half of the cases; 3) in these cases, the dissection of the bifurcation of the gastroduodenal artery and the vascular section beyond the origin of the infrapyloric artery allowed a direct vascular supply to the remaining duodenum to be preserved.This work was presented at the French Section of the European Association of Clinical Anatomy meeting, Bobigny, France, 1992     

Dithiothreitol prevents age-associated decrease in oocyte/conceptus viability in vitro

The present study was designed to ascertain whether the negative effects on reproductive potential of post-ovulatory ageing in vitro of oocytes can be prevented by antioxidant therapy. Mouse metaphase II (MII) oocytes were aged in vitro for 12 h prior to insemination in the presence of varying concentrations of L-ascorbic acid, 6-methoxy- 2,5,7,8-tetramethylchromane-2-carboxylic acid (Trolox), L-cystine dihydrochloride, ethylenediaminetetraacetic acid (EDTA), beta- mercaptoethanol and DL-dithiothreitol (DTT). In-vitro ageing of oocytes was associated with lower fertilization rate, higher proportion of concepti exhibiting cellular fragmentation at 24 h post-insemination and lower percentage of concepti reaching the blastocyst stage. Ascorbic acid, Trolox and EDTA had no effect on cellular fragmentation or potential of oocytes for development. However, the probability of an oocyte reaching the blastocyst stage was decreased (P < or = or = 0.05) in oocytes incubated in the presence of L-cystine (50 and 500 microM) and beta-mercaptoethanol (5, 50 and 500 microM) when compared to control aged oocytes. Age-associated cellular fragmentation at 24 h post-insemination was partially prevented (P < or = 0.05) by incubating oocytes in the presence of beta-mercaptoethanol (500 microM). DTT (50 and 500 microM) increased (P < or = 0.05) fertilization rate and number of cells at 81 h post-insemination to levels similar to those exhibited by control oocytes. Furthermore, both age-associated fragmentation at 24 h post-insemination (P < or = 0.05) and decreased potential of oocytes for development to the blastocyst stage (P < or = 0.05) were prevented, at least in part, by culturing oocytes in the presence of DTT (50 microM). Although the mechanism by which DTT exerts its beneficial effects on aged oocytes remains to be elucidated, it may protect oocytes by preventing oxidation of free thiol groups and/or altering a redox-independent signalling pathway that mediates cellular fragmentation and death.      

Characterization of 19 disease-associated missense mutations in the regulatory domain of the cystic fibrosis transmembrane conductance regulator

In order to gain a better insight into the structure and function of the regulatory domain (RD) of the cystic fibrosis transmembrane conductance regulator (CFTR) protein, 19 RD missense mutations that had been identified in patients were functionally characterized. Nine of these (I601F, L610S, A613T, D614G, I618T, L619S, H620P, G628R and L633P) resulted in aberrant processing. No or a very small number of functional CFTR proteins will therefore appear at the cell membrane in cells expressing these mutants. These mutations were clustered in the N- terminal part of the RD, suggesting that this subdomain has a folding pattern that is very sensitive to amino acid changes. Mutations that caused no aberrant processing were further characterized at the electrophysiological level. First, they were studied at the whole cell level in Xenopus laevis oocytes. Mutants that induced a whole cell current that was significantly different from wild-type CFTR were subsequently analysed at the single channel level in COS1 cells transiently expressing the different mutant and wild-type proteins. Three mutant chloride channels, G622D, R792G and E822K CFTR, were characterized by significantly lower intrinsic chloride channel activities compared with wild-type CFTR. Two mutations, H620Q and A800G, resulted in increased intrinsic chloride transport activities. Finally, T665S and E826K CFTR had single channel properties not significantly different from wild-type CFTR.