Critical illness is associated with insulin resistance and hyperglycemia. Intensive insulin treatment to normalize blood glucose during feeding has been shown to improve morbidity and mortality in ...patients in intensive care. The mechanisms behind the glucose-controlling effects of insulin in stress are not well understood.
Six previously healthy, severely traumatized patients (injury severity score > 15) were studied early (24–48 h) after trauma. Endogenous glucose production (EGP) and whole-body glucose disposal (WGD) were measured (6,6-2H2-glucose) at basal, during total parenteral nutrition (TPN), and during TPN plus insulin to normalize blood glucose (TPN+I). Six matched volunteers served as controls.
At basal and TPN, concentrations of glucose and insulin were higher in patients (P < 0.05). During TPN+I, insulin concentrations were 30-fold higher in patients. At basal, WGD and EGP were 30% higher in patients (P < 0.05). During TPN, EGP decreased in both groups but less in patients, resulting in 110% higher EGP than controls (P < 0.05). Normoglycemia coincided with reduced EGP, resulting in similar rates in both groups. WGD did not change during TPN or TPN+I and was not different between the groups.
In conclusion, in healthy subjects, euglycemia is maintained during TPN at physiological insulin concentrations by a reduction of EGP, whereas WGD is maintained at basal levels. In traumatized patients, hyperglycemia is due to increased EGP. In contrast to controls, normalization of glucose concentration during TPN needs high insulin infusion rates and is accounted for by a reduction in EGP, whereas WGD is not increased.
To examine the promoter methylation status of the 22 cancer genes and their contribution to disease progression in 6 head and neck squamous cell carcinoma (HNSCC) cell lines.
A panel of 41 gene ...probes, designed to interrogate 35 unique genes with known associations to cancer including HNSCC, was interrogated for alterations in gene copy number and aberrant methylation status (22 genes) using the methylation-specific multiplex ligation-dependent probe amplification assay.
Primary (A) and recurrent or metastatic (B) HNSCC cell lines UMSCC-11A/11B, UMSCC-17A/17B, and UMSCC-81A/81B are described.
Nine genes, TIMP3, APC, KLK10, TP73, CDH13, IGSF4, FHIT, ESR1, and DAPK1, were aberrantly methylated. The most frequently hypermethylated genes were APC and IGSF4, observed in 3 of 6 cell lines, and TP73 and DAPK1, observed in 2 of 6. For KLK10 and IGSF4, TIMP3 and FHIT, and TP73, in UMSCC-11B, UMSCC-17B, and UMSCC-81B, respectively, promoter hypermethylation was a disease progression event, indicating complete abrogation of tumor suppressor function for KLK10, IGSF4, and TIMP3 and gene silencing of 1 of 2 copies of TP73. Hypermethylation of IGSF4, TP73, CDH13, ESR1, DAPK1, and APC were primary events in UMSCC-17A.
Gene silencing through promoter hypermethylation was observed in 5 of 6 cell lines and contributed to primary and progressive events in HNSCC. In addition to genetic alterations of gains and losses, epigenetic events appear to further undermine a destabilized genomic repertoire in HNSCC.
We report on a fragile X mosaic male full mutation/normal allele detected by PCR and methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA). This combined analysis provides a ...diagnostic approach for fragile X syndrome (FXS). The method assesses the presence of expansion (full mutation), the CpG methylation status and could determine copy number changes (large deletions/duplications) along the FMR1 and FMR2 (fragile X mental retardation) genes. The method avoids detection of premutations, which makes it applicable for newborn screening. It can also be used in clarification of mosaic cases. The PCR results in our patient showed one normal allele; three repeats larger than his mother’s one. The MS-MLPA showed hypermethylated full mutation pattern in the proband. Both results are compatible with FXS mosaic case full mutation/normal allele. The patient demonstrates atypical mild clinical manifestation of the disease, which correlates to the presence of a normal size allele in the patient’s cells.
B cells use translesion DNA synthesis (TLS) to introduce somatic mutations around genetic lesions caused by activation-induced cytidine deaminase. Monoubiquitination at lysine(164) of proliferating ...cell nuclear antigen (PCNA(K164)) stimulates TLS. To determine the role of PCNA(K164) modifications in somatic hypermutation, PCNA(K164R) knock-in mice were generated. PCNA(K164R/K164R) mutants are born at a sub-Mendelian frequency. Although PCNA(K164R/K164R) B cells proliferate and class switch normally, the mutation spectrum of hypermutated immunoglobulin (Ig) genes alters dramatically. A strong reduction of mutations at template A/T is associated with a compensatory increase at G/C, which is a phenotype similar to polymerase eta (Poleta) and mismatch repair-deficient B cells. Mismatch recognition, monoubiquitinated PCNA, and Poleta likely cooperate in establishing mutations at template A/T during replication of Ig genes.
B cells use translesion DNA synthesis (TLS) to introduce somatic mutations around genetic lesions caused by activation-induced cytidine deaminase. Monoubiquitination at lysine super(164) of ...proliferating cell nuclear antigen (PCNA super(K164)) stimulates TLS. To determine the role of PCNA super(K164) modifications in somatic hypermutation, PCNA super(K164R) knock-in mice were generated. PCNA super(K164R/K164R) mutants are born at a sub-Mendelian frequency. Although PCNA super(K164R/K164R) B cells proliferate and class switch normally, the mutation spectrum of hypermutated immunoglobulin (Ig) genes alters dramatically. A strong reduction of mutations at template A/T is associated with a compensatory increase at G/C, which is a phenotype similar to polymerase eta (Pol eta ) and mismatch repair-deficient B cells. Mismatch recognition, monoubiquitinated PCNA, and Pol eta likely cooperate in establishing mutations at template A/T during replication of Ig genes.
Overnight fasting is routine before elective surgery. This may not be the optimal way to prepare for surgical stress, however, because intravenous carbohydrate supplementation instead of fasting has ...recently been shown to reduce postoperative insulin resistance. In the current study, gastric emptying of a carbohydrate-rich drink was investigated before elective surgery and in a control situation.
Twelve patients scheduled for elective surgery were randomly given 400 mL of either a carbohydrate-rich drink (285 mOsm/kg, 12.0% carbohydrates, n = 6) or water 4 hours before being anesthetized. Gastric emptying was measured (gamma camera, 99Tcm). Each patient repeated the protocol postoperatively as a control. All values were presented as the mean +/- SEM by means of a nonparametric statistical evaluation.
Despite the increased anxiety experienced by patients before surgery (p < 0.005), gastric emptying did not differ between the experimental and control situations. Initially, water emptied more rapidly than carbohydrate. However, after 90 minutes, the stomach was emptied regardless of the solution administered (3.2 +/- 1.1% mean +/- SEM remaining in the stomach in the carbohydrate group versus 2.3 +/- 1.2% remaining in the stomach in the water group).
Preoperative anxiety does not prolong gastric emptying. The stomach had been emptied 90 minutes after ingestion of both the carbohydrate-rick drink and water, thereby indicating the possibility of allowing an intake of iso-osmolar carbohydrate-rich fluids before surgery.
Large deletions in the APC (adenomatous polyposis coli) gene, causing familial adenomatous polyposis (FAP), cannot easily be detected by conventional mutation-detection techniques. Therefore, we have ...developed two independent quantitative methods for the detection of large deletions, encompassing one or more exons, of APC. Multiplex ligation-dependent probe amplification (MLPA) is performed in one reaction for the initial quantification of all APC exon copy numbers. Subsequently, quantitative real-time PCR (QRT-PCR) is used to verify the results obtained in the MLPA reaction. The identification of a deletion of the whole APC gene in a patient with classical FAP is described. The mutation was detected with the two quantitative methods and further verified on chromosomal level by the use of FISH (fluorescence in situ hybridization) on metaphase spreads. Furthermore, a large deletion covering exons 11-13 of the APC gene was detected in two apparently unrelated families. This deletion was further verified and characterized with long-range PCR. The MLPA test ensures a sensitive high-throughput screening for large deletions of the APC gene and can easily be implemented in the diagnostic testing for FAP.
A consortium of seven European partners has undertaken the challenging task to develop a miniaturized system capable to isolate and characterize circulating tumor cells (CTC), which should have its ...use as a time and labor saving, minimally invasive tool for cancer therapy selection and monitoring and for the detection of recurrent disease. This paper reports on the general concept of the system and elaborates on the development of the different modules and their integration.
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