Measurement of estradiol-17-fatty acid esters in human tissues.

We have developed a gas chromatographic/mass spectral method for the sensitive and reproducible measurement of estradiol-17-fatty acid esters in human tissues and blood. To provide an internal standard for quantification, a trideuterated analog of a representative estradiol ester is added to the tissues. Estradiol (E2) released from the nonpolar ester fraction by alkaline hydrolysis is derivatized to form the ditrimethylsilyl ether and then analyzed by gas chromatographic/mass spectral, monitoring the molecular ions mass per U charge of the ditrimethylsilyl derivative of E2 and [2H3]E2. There are low but detectable levels of E2 ester in the blood of cycling females; there are none in urine. While the E2 ester is present in breast cyst fluid, its concentration, 77-140 pmol/L, is considerably less than E2, 110-2,863 pmol/L. But there is a large amount of E2 ester in fat. In premenopausal women the average E2 ester in fat (sc and omental) is 957 +/- 283 38 fmol/g (SEM); in women who are menopausal less than 12 yr, the E2 ester in fat is 669 +/- 158 fmol/g; in women who are menopausal at least 15 yr, the fat level is 399 +/- 146 fmol/g. Muscle from the same women have lower concentrations of the ester; in 8 out of 12 muscle specimens it was not detectable. The E2 esters are extremely potent estrogens. Although they are hormonally active they require enzymatic hydrolysis to exert their hormonal action. These studies show that these long chain esters of E2 are sequestered in fatty tissues, wherein they represent a protected store of preformed hormone. Under the proper stimulation, adipose tissue can activate the estrogenic signal through the action of hormonally sensitive esterases. Thus, through signaling between estrogen sensitive tissues and neighboring fat cells, a local paracrine loop may exist.     

Detection of chromosomes and estimation of aneuploidy in human spermatozoa using fluorescence in-situ hybridization

The development and application of fluorescence in-situ hybridization (FISH) has opened the way for comprehensive studies on numerical chromosome abnormalities in human spermatozoa. FISH can be rapidly applied to large numbers of spermatozoa and thus overcomes the major limitation of karyotyping spermatozoa after penetration of zona-free hamster oocytes. The simultaneous hybridization of two or more chromosome-specific probes to spermatozoa and subsequent detection of the bound probes using different fluorescent detection systems enables two or more chromosomes to be localized simultaneously in the same spermatozoon and provides a technique for undertaking reasonable estimates of aneuploidy. The most commonly used probes are those which bind to the centromeric region of specific chromosomes. Most studies to date have concentrated on estimating aneuploidy in spermatozoa from normospermic men, although reports are beginning to appear on aneuploidy in spermatozoa from subfertile and infertile men. Multi- probe FISH studies have generally reported disomy (hyperhaploidy) estimates of 0.05-0.2% per chromosome. There is preliminary evidence that some chromosomes such as X, Y and 21 are predisposed towards higher rates of non-disjunction during spermatogenesis. There are also suggestions of inter-donor variability in aneuploidy frequencies for specific chromosomes, although this requires confirmation in larger studies. While FISH is clearly a powerful technique that has many applications in reproductive medicine, it must also be realized that it does have limitations and the technology itself is still evolving and has yet to be fully validated on spermatozoa.      

Identification of MEN1 gene mutations in sporadic carcinoid tumors of the lung

Lung carcinoids occur sporadically and rarely in association with multiple endocrine neoplasia type 1 (MEN1). There are no well defined genetic abnormalities known to occur in these tumors. We studied 11 sporadic lung carcinoids for loss of heterozygosity (LOH) at the locus of the MEN1 gene on chromosome 11q13, and for mutations of the MEN1 gene using dideoxy fingerprinting. Additionally, a lung carcinoid from a MEN1 patient was studied. In four of 11 (36%) sporadic tumors, both copies of the MEN1 gene were inactivated. All four tumors showed the presence of a MEN1 gene mutation and loss of the other allele. Observed mutations included a 1 bp insertion, a 1 bp deletion, a 13 bp deletion and a single nucleotide substitution affecting a donor splice site. Each mutation predicts truncation or potentially complete loss of menin. The remaining seven tumors showed neither the presence of a MEN1 gene mutation nor 11q13 LOH. The tumor from the MEN1 patient showed LOH at chromosome 11q13 and a complex germline MEN1 gene mutation. The data implicate the MEN1 gene in the pathogenesis of sporadic lung carcinoids, representing the first defined genetic alteration in these tumors.      

Problem solving,sense of coherence and instrumental ADL of elderly people with depression and normal control group

The purpose of this study was first, to explore the separate contribution and interaction between verbal and performance based problem solving and sense of coherence; and second, to examine the association of these variables with Instrumental Activities of Daily Living (IADL) function among elderly people with depression living in the commuinity and a normal control group. The participants included elders receiving ambulatory care for depression (n=31; mean age=73, SD=9.3); and normal elders (n=30; mean age=78, SD=5.8). Screening for general cognitive ability and level of depression was done using the Mini Mental Status Examination (MMSE) and the Geriatric Depression Scale (GDS). All participants underwent evaluation using the Large Allen Cognitive Level Test (LACL) (a measure of performance based problem solving); the Problem Solving Verbal Reaction to everyday problematic situations (PSVR); the Sense of Coherence questionnaire (SOC) and the Routine Task Inventory (RTI) (a measure of IADL from the cognitive perspective). Wilcoxon statistical analysis indicated highly significant differences between the two groups for all of the study variables. In the depressive group, significant correlations were found between the components of problem solving (LACL and PSVR) and IADL (r=0.70 and r=0.53), while the SOC did not correlate with IADL in either group. Results of ANCOVA controlling for LACL showed that it has a significant effect (F=13.63, p=0.001); however, beyond it verbal problem solving still has a significant effect on IADL (F=4.77, p=0.02), and SOC in interaction with verbal problem solving was significant (F=3.97, p=0.035). The findings suggest that depression in elderly people is associated with lower functioning in problem solving and IADL function, and lower sense of coherence; hence, attention to these factors should be integral to intervention with elderly people. However, it is recommended that further study be made of the relationships of variables found in this study with the current instruments, and also with additional tools because of confounding effects, in order to further support and validate the findings. As the sample size was small compared with the number of measures, it is important to replicate the study with larger groups to have more power. Copyright © 1999 Whurr Publishers Ltd.     

Preoperative spirometry predicts perioperative pulmonary complications after major vascular surgery.

This study determined utility of preoperative spirometry for prediction of postoperative pulmonary complications (PPC) defined as pneumonia, ventilator dependence greater than 48 hours, and adult respiratory distress syndrome in 147 patients undergoing vascular surgery from June 1988 through March 1990 [39 aortic aneurysm repairs, 21 carotid procedures, and 87 operations for occlusive disease including aorto-ileofemoral, infra-inguinal, and visceral]. The incidence of PPC was 12.9 per cent, while cardiac complications (myocardial infarction, congestive heart failure, and ventricular arrhythmias) were present in 9.8 per cent. Prior or current smoking, which was present in 80 per cent, was not predictive of PPC. FEV1 was 2.2 +/- 0.7 L/s (mean +/- 1 SD). Abnormal FEV1 (2.0 or less L/s) was present in 42 per cent (n = 62). For FEV1 of 2.0 or less, PPC rate was 22.5 per cent versus 5.8 per cent for FEV1 greater than 2.0 L/s (P less than 0.005, Fisher exact). The incidence of PPC was 30.7 per cent for aortic aneurysm repair, 8.0 per cent for occlusive disease, and 4.7 per cent for carotid procedures. Abdominal aortic procedures (performed in 67 patients: 39 for aortic aneurysm repair and 28 for aortoiliac occlusive disease) were associated with a PPC rate of 22.4 per cent versus 5.0 per cent for "nonabdominal" procedures (P less than 0.002, Fisher exact). Life table analysis after surgery demonstrated decreased survival for patients with PPC (P = 0.031, Mantel-Haensel) during follow-up (250 +/- 165 days). PPC are associated with abnormal FEV1 and abdominal vascular procedures. In conclusion, preoperative spirometry is useful for the prediction of PPC after vascular surgery.     

A prospective study of methylenetetrahydrofolate reductase and methionine synthase gene polymorphisms, and risk of colorectal adenoma

We examined the relationship between a functional polymorphism (667C-- >T, ala-->val) of the methylenetetrahydrofolate reductase gene (MTHFR) and the risk of colorectal adenomas in the prospective Nurses' Health Study. Among 257 incident polyp cases and 713 controls, the MTHFR val/val polymorphism [relative risk (RR) = 1.35, 95% confidence interval (CI) 0.84-2.17] was not significantly associated with risk of adenomas. This lack of association was observed for both small (RR = 1.36, 95% CI 0.76-2.45) and large (RR = 1.32, 95% CI 0.66-2.66) adenomas. Furthermore, there was no significant interaction between this polymorphism and consumption of either folate, methionine or alcohol. We also examined the relationship of a newly identified polymorphism (asp919gly) of the methionine synthase gene (MS) with the risk of colorectal adenomas in the same population. The MS gly/gly polymorphism was also not significantly associated with risk of colorectal adenomas (RR = 0.66, 95% CI 0.26-1.70). These results, which need to be confirmed in other studies, suggest that the MTHFR val/val polymorphism, which has been previously inversely associated with risk of colorectal cancer, plays a role only in a late stage (adenoma-- >carcinoma) of colorectal tumorigenesis, and/or may protect against malignant transformation in the subset of benign adenomas, which may progress to malignancy.      

Quercetin alters the DNA damage response in human hematopoietic stem and progenitor cells via TopoII‐ and PI3K‐dependent mechanisms synergizing in leukemogenic rearrangements

Quercetin (Que) is an abundant flavonoid in the human diet and high‐concentration food supplement with reported pro‐ and anti‐carcinogenic activities. Topoisomerase II (TopoII) inhibition and subsequent DNA damage induction by Que was implicated in the mixed lineage leukemia gene (MLL) rearrangements that can induce infant and adult leukemias. This notion raised concerns regarding possible genotoxicities of Que in hematopoietic stem and progenitor cells (HSPCs). However, molecular targets mediating Que effects on DNA repair relevant to MLL translocations have not been defined. In this study we describe novel and potentially genotoxic Que activities in suppressing non‐homologous end joining and homologous recombination pathways downstream of MLL cleavage. Using pharmacological dissection of DNA‐PK, ATM and PI3K signalling we defined PI3K inhibition by Que with a concomitant decrease in the abundance of key DNA repair genes to be responsible for DNA repair inhibition. Evidence for the downstream TopoII‐independent mutagenic potential of Que was obtained by documenting further increased frequencies of MLL rearrangements in human HSPCs concomitantly treated with Etoposide and Que versus single treatments. Importantly, by engaging a tissue engineered placental barrier, we have established the extent of Que transplacental transfer and hence provided the evidence for Que reaching fetal HSPCs. Thus, Que exhibits genotoxic effects in human HSPCs via different mechanisms when applied continuously and at high concentrations. In light of the demonstrated Que transfer to the fetal compartment our findings are key to understanding the mechanisms underlying infant leukemia and provide molecular markers for the development of safety values.