Linkage disequilibrium studies in multiple endocrine neoplasia type 1 (MEN1)

Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant disorder characterised by tumours of the parathyroids, pancreas and anterior pituitary. The MEN1 gene has been localised to a 2-Mb region of chromosome 11q13 by meiotic mapping studies in MEN1 families. Such studies may have a limited resolution of approximately 1 cM (i.e. 1 Mb) and we have therefore investigated 96 MEN1 families (40 British, 17 French, 12 Finnish, 7 Swedish, 7 Dutch, 7 North American, 2 Australian, 1 New Zealand, 1 German, 1 Spanish and 1 Danish) for linkage disequilibrium, in order to facilitate a finer mapping resolution. We have utilised five microsatellite DNA sequence polymorphisms from the candidate region and have accurately determined their allele sizes, which ranged from 161 bp to 272 bp. The heterozygosity and number of alleles (given in brackets), respectively, at the loci were: D11S1883 (76%, 11), D11S457 (55%, 5), PYGM (94%, 18), D11S1783 (10%, 4) and D11S449 (87%, 16). Allelic association was assessed by Chi-square 2 ×n contingency tables, by Fisher exact 2 ×n contingency tables and by a likelihood-based approach. The results of haplotype analysis revealed 91 different affected haplotypes in the 96 families, an identical affected haplotype being observed in no more than two families. These results indicate the absence of an ancestral affected haplotype. Significant linkage disequilibrium (P < 0.005) could be established amongst the microsatellite loci but not between the loci and MEN1 in either the total population or in any of the geographical sub-populations. The absence of linkage disequilibrium between MEN1 and the polymorphic loci is probably the result of the occurrence of multiple different disease-causing mutations in MEN1. Received: 1 April 1997 / Accepted: 25 June 1997     

Effect of medium composition, agitation and the presence of EDTA on the antimicrobial activity of cryptolepine

The antimicrobial activity of cryptolepine is influenced by the type of medium employed, agitation and the presence of non-inhibitory concentrations of EDTA. The use of Mueller–Hinton broth (MHB), iso-sensitest broth and tryptone soya broth (TSB) produced lower minimum inhibitory concentrations (MICs) for some of the test organisms compared with nutrient broth or yeast dextrose broth (YDB). For example, a fourfold drop in MIC was recorded for Saccharomyces cerevisiae in MHB compared with the same organism tested in YDB. Agitation of the broths during incubation nearly always produced lower MICs for the bacteria, an eightfold decrease in MIC being recorded for Escherichia coli cultured in nutrient broth with agitation compared with a statically maintained culture. A non-inhibitory concentration (10⁻³ mol l⁻¹) of disodium EDTA enhanced the antimicrobial activity of cryptolepine. Against E. coli NCTC 11560, an eightfold decrease in MIC and minimum bactericidal concentration (MBC) was recorded when tested in the presence of EDTA.     

Evidence of differential renal dysfunctions during exercise in men

Post-exercise proteinuria is a common phenomenon in healthy subjects. Previous studies have used albumin (Alb) and β₂-microglobulin (β₂-m) molecules as representatives of high- and low-molecular-weight proteins. Recently, more specific markers of the human kidney proximal tubule have been used to identify the precise site of alterations. Active male subjects underwent two strenuous runs, one 400-m run and one 3000-m run. Urine was collected from the subjects before and after each event. Total protein (TP), Alb, α₁-microglobulin (α₁-m), β₂-m, intestinal alkaline phosphatase (IAP), tissue-nonspecific alkaline phosphatase (TNAP) and N-acetyl-β-d-glucosaminidase (NAG) were determined for each sample. The short-distance run (400 m) resulted in the largest increases (P ≤ 0.05) in TP (31-fold), Alb (100-fold) and β₂-m (164-fold) as compared to the long-distance run (3000-m). The α₁-m excretion rates were increased to a lesser extent by the exercises. The IAP activity was slightly increased (+90%) by the 400-m run while the TNAP and NAG activities showed a 6.8-fold and a 3.6-fold increase, respectively, after this event. Smaller increases were recorded for the long-distance run (P = 0.05). To conclude, the present investigation showed that: (1) post-exercise proteinuria is related to the absolute intensity of exercise; (2) the impairment of protein reabsorption is revealed better by changes in Alb and β₂-m; (3) changes in TNAP and NAG activities could reveal biochemical modifications that occur in the proximal tubule, particularly at the S1-S2 segment. Accepted: 31 January 1997     

Mitochondrial function in oncogene-transfected rat fibroblasts isolated from multicellular spheroids

Twomitochondrion-specific fluorochromes,10-N-nonyl acridineorange (NAO) and rhodamine 123 (Rh123), were used todetermine the mechanism responsible for alterations in energymetabolism of transformed rat embryo fibroblast cells isolated fromdifferent locations within multicellular spheroids. Accumulation ofRh123 depends on intact mitochondrial membrane potential, whereas NAO is taken up by mitochondria independently of their function and thusrepresents mitochondrial distribution only. A reproducible selectivedissociation procedure was used to isolate cells from differentlocations within the spheroids. After isolation, cells weresimultaneously stained with one mitochondrial stain and the DNA dyeHoechst 33342, and several parameters, including cell volume, weremonitored via multilaser-multiparameter flow cytometry. Our dataclearly show a decrease in the uptake of Rh123 in cells from theperiphery to the inner regions of the tumor spheroids, reflecting apersistent alteration in mitochondrial function. However, NAO stainingexperiments showed no reduction in the total mitochondrial mass perunit cell volume. Because cells were exposed to stain under uniformconditions after isolation from the spheroid, these data indicate thatdownregulation of mitochondrial function is associated with cellquiescence rather than a transient effect of reduced nutrientavailability. This result, which is in accordance with data from twoother cell lines (EMT6 and 9L), might reflect a general phenomenon inmulticellular spheroids, supporting the hypothesis that quiescent cellsin the innermost viable spheroid layer stably reduce theirmitochondrial function, presumably to compensate for lower nutrientsupply and/or decreased energy demand.