Morphological analysis of degeneration and regeneration of syncytiotrophoblast in first trimester placental villi during organ culture

We have recently shown using dansyl-L-lysine exclusion studies that the release of human chorionic gonadotrophin (HCG) in conjunction with L- lactate dehydrogenase (LDH) from first trimester villi during organ culture is symptomatic of syncytiotrophoblast degeneration. The purpose of this study was to examine chorionic villi at the ultrastructural level in order to determine events occurring during organ culture. The tissue was sampled after 0, 24, 48 and 120 h in culture and processed for electron microscopy. In addition to confirming the previously recorded syncytial degeneration, the electron micrographs showed clearly the generation of a new syncytiotrophoblast layer. The new layer, derived from differentiating cytotrophoblast cells, was largely formed by 48 h and was maintained for at least 120 h in culture. This study demonstrates a model which provides an opportunity to study the differentiation of cytotrophoblast cells whilst they retain their anatomical relationships within the villous structure.      

Insulin and epinephrine effects on heart glycogen synthase and phosphorylase activity.

The effect of intravenous epinephrine on heart glycogen synthase and phosphorylase systems in control and insulin-pretreated rats was studied. The percent of synthase in the I form decreased rapidly after epinephrine treatment but the change was small and sometimes not significant. In insulin-pretreated rats in which the percent synthase I was increased, epinephrine produced a definate and highly significant decrease. There was a simultaneous increase in percent phosphorylase a in both groups. The synthase and phosphorylase responses were statiscally significant at 2.5 mug epinephrine/kgor more. These data are compatible with a mechanism in which protein kinase is activated by an increased cAMP concentration and affects both the synthase and phosphorylasesystems simultaneously. Propranolol blocked the epinephrine effects on cAMP, synthase I, and phosphorylase a. Although insulin had little effect on the response ofthe synthase and phosphorylase systems to epinephrine, it nealry completely blocked glycogen degradation. The mechanism is unknown, but it appears to be due to an inhibition of phosphorylase a catalytic activity in vivo. Acetylcholine had no effect on synthase I, phosphorylase a, or cAMP in control or in insulin-pretreated animals.     

Mutation spectrum and genotype-phenotype analyses in Cowden disease and Bannayan-Zonana syndrome, two hamartoma syndromes with germline PTEN mutation

The tumour suppressor gene PTEN , which maps to 10q23.3 and encodes a 403 amino acid dual specificity phosphatase (protein tyrosine phosphatase; PTPase), was shown recently to play a broad role in human malignancy. Somatic PTEN deletions and mutations were observed in sporadic breast, brain, prostate and kidney cancer cell lines and in several primary tumours such as endometrial carcinomas, malignant melanoma and thyroid tumours. In addition, PTEN was identified as the susceptibility gene for two hamartoma syndromes: Cowden disease (CD; MIM 158350) and Bannayan-Zonana (BZS) or Ruvalcaba-Riley-Smith syndrome (MIM 153480). Constitutive DNA from 37 CD families and seven BZS families was screened for germline PTEN mutations. PTEN mutations were identified in 30 of 37 (81%) CD families, including missense and nonsense point mutations, deletions, insertions, a deletion/insertion and splice site mutations. These mutations were scattered over the entire length of PTEN , with the exception of the first, fourth and last exons. A 'hot spot' for PTEN mutation in CD was identified in exon 5 that contains the PTPase core motif, with 13 of 30 (43%) CD mutations identified in this exon. Seven of 30 (23%) were within the core motif, the majority (five of seven) of which were missense mutations, possibly pointing to the functional significance of this region. Germline PTEN mutations were identified in four of seven (57%) BZS families studied. Interestingly, none of these mutations was observed in the PTPase core motif. It is also worthy of note that a single nonsense point mutation, R233X, was observed in the germline DNA from two unrelated CD families and one BZS family. Genotype-phenotype studies were not performed on this small group of BZS families. However, genotype-phenotype analysis inthe group of CD families revealed two possible associations worthy of follow-up in independent analyses. The first was an association noted in the group of CD families with breast disease. A correlation was observed between the presence/absence of a PTEN mutation and the type of breast involvement (unaffected versus benign versus malignant). Specifically and more directly, an association was also observed between the presence of a PTEN mutation and malignant breast disease. Secondly, there appeared to be an interdependent association between mutations upstream and within the PTPase core motif, the core motif containing the majority of missense mutations, and the involvement of all major organ systems (central nervous system, thyroid, breast, skin and gastrointestinal tract). However, these observations would need to be confirmed by studying a larger number of CD families.      

A dinucleotide mutation in the endothelin-B receptor gene is associated with lethal white foal syndrome (LWFS); a horse variant of Hirschsprung disease

Lethal white foal syndrome (LWFS) is a congenital anomaly of horses characterized by a white coat colour and aganglionosis of the bowel, which is similar to Hirschsprung disease (HSCR). We decided to investigate possible mutations of the endothelin-B receptor gene ( EDNRB ) in LWFS as recent studies in mutant rodents and some patients have demonstrated EDNRB defects. First, we identified a full-length cDNA for horse EDNRB . This cDNA fragment contained a 1329 bp open reading frame which encoded 443 amino acid residues. The predicted amino acid sequence was 89, 91 and 85% identical to human, bovine and mouse as well as rat EDNRB respectively, but only 55% identical to the human, bovine and rat endothelin A receptor (EDNRA). Secondly, sequence analysis, together with allele-specific PCR and the amplification- created restriction site (ACRS) technique, revealed a dinucleotide TC-- >AG mutation, which changed isoleucine to lysine in the predicted first transmembrane domain of the EDNRB protein. This was associated with LWFS when homozygous and with the overo phenotype when heterozygous.      

Maternal serum alpha-fetoprotein and altitude

Maternal serum alpha-fetoprotein (MS-alphaFP) testing is widely used to screen for fetal defects. MS-alphaFP concentrations are affected by a number of variables such as gestational age, maternal weight, number of fetuses, race, and insulin-dependent diabetes. Undefined geographic factors may also influence MS-alphaFP. We have examined the effect of altitude in a sample of 1063 MS-alphaFP results selected to span a range of altitudes. The study sample was subjected to linear regression with and without a term for altitude, and multiple-of-the-median (MoM) values were calculated before and after adjusting for altitude. The median MS-alphaFP was found to decrease an average of 1 ng/mL for every 1100 ft increase in altitude, a change approximately equivalent to that seen with an increase in maternal weight of 6 lb. Adjusting for altitude resulted in the reclassification of 36 of 1063 patient results (3.4%), although the clinical utility of this adjustment remains unexamined.